Nucleotide sequences which code for the rpoB gene

ABSTRACT

The present invention relates to polynucleotides corresponding to the rpoB gene and which encode the β-subunit of RNA polymerase B, methods of producing L-amino acids, and methods of screening for polynucleotides which encode proteins having activity of the β-subunit of RNA polymerase B.

CROSS-REFERENCE TO RELATED APPLICATION

[0001] The present application claims priority to German Application No. DE10107229.5 filed Feb. 16, 2001, the entire contents of which are incorporated herein by reference.

BACKGROUND OF THE INVENTION

[0002] 1. Field of the Invention

[0003] The present invention relates to polynucleotides corresponding to the rpoB gene and which encode the β-subunit of RNA polymerase B, methods of producing L-amino acids, and methods of screening for polynucleotides which encode proteins having activity of the β-subunit of RNA polymerase B.

[0004] 2. Discussion of the Background

[0005] L-amino acids, especially L-lysine, are used in human medicine and in the pharmaceuticals industry, in the foodstuffs industry and, very especially, in the feeding of animals.

[0006] It is known that amino acids are produced by fermentation of strains of coryneform bacteria, especially Corynebacterium glutamicum. Because of their great importance, attempts are continuously being made to improve the production processes. Improvements to the processes may concern measures relating to the fermentation, such as, for example, stirring and oxygen supply, or the composition of the nutrient media, such as, for example, the sugar concentration during the fermentation, or working up to the product form by, for example, ion-exchange chromatography, or the intrinsic performance properties of the microorganism itself.

[0007] In order to improve the performance properties of such microorganisms, methods of mutagenesis, selection and mutant selection are employed. Such methods yield strains which are resistant to antimetabolites or are auxotrophic for metabolites that are important in terms of regulation, and which produce amino acids.

[0008] For a number of years, methods of recombinant DNA technology have also been used for improving the strain of L-amino acid-producing strains of Corynebacterium, by amplifying individual amino acid biosynthesis genes and studying the effect on amino acid production.

[0009] However, there remains a critical need for improved methods of producing L-amino acids and thus for the provision of strains of bacteria producing higher amounts of L-amino acids. On a commercial or industrial scale even small improvements in the yield of L-amino acids, or the efficiency of their production, are economically significant. Prior to the present invention, it was not recognized that enhancement of the rpoB gene encoding the β-subunit of RNA polymerase B would improve L-amino acid yields.

SUMMARY OF THE INVENTION

[0010] One object of the present invention, is providing a new process adjuvant for improving the fermentative production of L-amino acids, particularly L-lysine and L-glutamate. Such process adjuvants include enhanced bacteria, preferably enhanced Coryneform bacteria which express enhanced levels of the β-subunit of RNA polymerase B which is encoded by the rpoB gene.

[0011] Thus, another object of the present invention is providing such a bacterium, which expresses enhanced amounts of the β-subunit of RNA polymerase B or gene products of the rpoB gene.

[0012] Another object of the present invention is providing a bacterium, preferably a Coryneform bacterium, which expresses a polypeptide that has an enhanced β-subunit of RNA polymerase B activity.

[0013] Another object of the invention is to provide a nucleotide sequence encoding a polypeptide which has a β-subunit of RNA polymerase B sequence. One embodiment of such a sequence is the nucleotide sequence of SEQ ID NO: 1. Other embodiments of such a sequence is the nucleotide sequences of SEQ ID NOS:3 and 5.

[0014] A further object of the invention is a method of making a β-subunit of RNA polymerase B or an isolated polypeptide having a β-subunit of RNA polymerase B activity, as well as use of such isolated polypeptides in the production of amino acids. One embodiment of such a polypeptide is the polypeptide having the amino acid sequence of SEQ ID NO: 2. Other embodiments of such a sequence is the amino acid sequence of SEQ ID NOS:4 and 6.

[0015] In one embodiment the invention provides isolated polypeptides comprising the amino acid sequences in SEQ ID NOS:2, 4 and/or 6.

[0016] Other objects of the invention include methods of detecting nucleic acid sequences homologous to SEQ ID NO: 1, particularly nucleic acid sequences encoding polypeptides that have the activity of a β-subunit of RNA polymerase B, and methods of making nucleic acids encoding such polypeptides.

[0017] The above objects highlight certain aspects of the invention. Additional objects, aspects and embodiments of the invention are found in the following detailed description of the invention.

DETAILED DESCRIPTION OF THE INVENTION

[0018] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art of molecular biology. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described herein. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and are not intended to be limiting.

[0019] Reference is made to standard textbooks of molecular biology that contain definitions and methods and means for carrying out basic techniques, encompassed by the present invention. See, for example, Maniatis et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, New York (1982) and Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, New York (1989) and the various references cited therein.

[0020] Where L-amino acids or amino acids are mentioned hereinbelow, they are to be understood as meaning one or more amino acids, including their salts, selected from the group L-asparagine, L-threonine, L-serine, L-glutamate, L-glycine, L-alanine, L-cysteine, L-valine, L-methionine, L-isoleucine, L-leucine, L-tyrosine, L-phenylalanine, L-histidine, L-lysine, L-tryptophan and L-arginine. L-lysine is especially preferred.

[0021] Where L-lysine or lysine is mentioned hereinbelow, it is to be understood as meaning not only the bases but also the salts, such as, for example, lysine monohydrochloride or lysine sulfate.

[0022] The invention provides an isolated polynucleotide from coryneform bacteria, containing a polynucleotide sequence coding for the rpoB gene, selected from the group

[0023] a) polynucleotide that is at least 70% identical with a polynucleotide that codes for a polypeptide containing the amino acid sequence of SEQ ID No. 2,

[0024] b) polynucleotide that codes for a polypeptide containing an amino acid sequence that is at least 70% identical with the amino acid sequence of SEQ ID No. 2,

[0025] c) polynucleotide that is complementary to the polynucleotides of a) or b), and

[0026] d) polynucleotide containing at least 15 consecutive nucleotides of the polynucleotide sequence of a), b) or c),

[0027] the polypeptide preferably exhibiting the activity of the β-subunit of RNA polymerase B.

[0028] The invention also provides the above-mentioned polynucleotide, it preferably being a replicatable DNA containing:

[0029] (i) the nucleotide sequence shown in SEQ ID No. 1, or

[0030] (ii) at least one sequence that corresponds to sequence (i) within the region of the degeneracy of the genetic code, or

[0031] (iii) at least one sequence that hybridizes with the sequence that is complementary to sequence (i) or (ii), and optionally

[0032] (iv) sense mutations in (i) which are neutral in terms of function and which do not change the activity of the protein/polypeptide.

[0033] Finally, the invention also provides polynucleotides selected from the group

[0034] a) polynucleotides containing at least 15 consecutive nucleotides selected from the nucleotide sequence of SEQ ID No. 1 between positions 1 and 701

[0035] b) polynucleotides containing at least 15 consecutive nucleotides selected from the nucleotide sequence of SEQ ID No. 1 between positions 702 and 4199

[0036] c) polynucleotides containing at least 15 consecutive nucleotides selected from the nucleotide sequence of SEQ ID No. 1 between positions 4200 and 5099.

[0037] The invention also provides

[0038] a replicatable polynucleotide, especially DNA, containing the nucleotide sequence as shown in SEQ ID No. 1;

[0039] a polynucleotide that codes for a polypeptide containing the amino acid sequence as shown in SEQ ID No. 2;

[0040] a vector containing the polynucleotide of the invention, especially a shuttle vector or a plasmid vector, and

[0041] coryneform bacteria which contain the vector or in which the rpoB gene has been enhanced.

[0042] The invention also provides polynucleotides consisting substantially of a polynucleotide sequence, which are obtainable by screening, by means of hybridization, a corresponding gene library of a coryneform bacteria that contains the complete gene or parts thereof, using a probe containing the sequence of the polynucleotide of the invention according to SEQ ID No. 1 or a fragment thereof, and isolating the mentioned polynucleotide sequence.

[0043] Polynucleotides that contain the sequences of the invention are suitable as hybridization probes for RNA, cDNA and DNA, in order to isolate in their complete length nucleic acids or polynucleotides or genes that code for the β-subunit of RNA polymerase B, or in order to isolate nucleic acids or polynucleotides or genes that are very similar to the sequence of the rpoB gene. They are likewise suitable for incorporation into so-called “arrays”, “micro arrays” or “DNA chips” in order to detect and determine the corresponding polynucleotides.

[0044] Polynucleotides that contain the sequences of the invention are also suitable as primers, with the aid of which it is possible, by means of the polymerase chain reaction (PCR), to produce DNA of genes that code for the β-subunit of RNA polymerase B.

[0045] Such oligonucleotides acting as probes or primers contain at least 25, 26, 27, 28, 29 or 30, preferably at least 20, 21, 22, 23 or 24, very especially preferably at least 15, 16, 17, 18 or 19, consecutive nucleotides. Also suitable are oligonucleotides having a length of at least 31, 32, 33, 34, 35, 36, 37, 38, 39 or 40 or of at least 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 nucleotides. Oligonucleotides having a length of at least 100, 150, 200, 250 or 300 nucleotides may also be suitable.

[0046] “Isolated” means removed from its natural environment.

[0047] “Polynucleotide” generally refers to polyribonucleotides and polydeoxyribonucleotides, it being possible for the RNA or DNA to be unmodified or modified.

[0048] The polynucleotides of the invention include a polynucleotide according to SEQ ID No. 1 or a fragment prepared therefrom, and also polynucleotides that are at least especially from 70% to 80%, preferably at least from 81% to 85%, especially preferably at least from 86% to 90%, and very especially preferably at least 91%, 93%, 95%, 97% or 99%, identical with the polynucleotide according to SEQ ID No. 1, or with a fragment prepared therefrom.

[0049] “Polypeptides” are to be understood as being peptides or proteins that contain two or more amino acids bonded via peptide bonds.

[0050] The polypeptides of the invention include a polypeptide according to SEQ ID No. 2, especially those having the biological activity of the β-subunit of RNA polymerase B, and also those that are at least from 70% to 80%, preferably at least from 81% to 85%, especially preferably at least from 86% to 90%, and very especially preferably at least 91%, 93%, 95%, 97% or 99%, identical with the polypeptide according to SEQ ID No. 2 and exhibit the mentioned activity.

[0051] The invention also provides a process for the production, by fermentation, of amino acids selected from the group L-asparagine, L-threonine, L-serine, L-glutamate, L-glycine, L-alanine, L-cysteine, L-valine, L-methionine, L-isoleucine, L-leucine, L-tyrosine, L-phenylalanine, L-histidine, L-lysine, L-tryptophan and L-arginine, using coryneform bacteria which, in particular, already produce amino acids and in which the nucleotide sequences coding for the rpoB gene are enhanced, especially overexpressed.

[0052] The term “enhancement” in this connection describes the increasing of the intracellular activity of one or more enzymes or proteins in a microorganism that are coded for by the corresponding DNA, by, for example, increasing the number of copies of the gene or genes, using a strong promoter or using a gene or allele that codes for a corresponding enzyme or protein having a high level of activity, and optionally by combining those measures.

[0053] The microorganisms provided by the present invention can produce L-amino acids from glucose, saccharose, lactose, fructose, maltose, molasses, starch, cellulose or from glycerol and ethanol. They may be representatives of coryneform bacteria, especially of the genus Corynebacterium. In the case of the genus Corynebacterium, special mention may be made of the species Corynebacterium glutamicum, which is known to those skilled in the art for its ability to produce L-amino acids.

[0054] Suitable strains of the genus Corynebacterium, especially of the species Corynebacterium glutamicum (C. glutamicum), are especially the known wild-type strains

[0055]Corynebacterium glutamicum ATCC13032

[0056]Corynebacterium acetoglutamicum ATCC15806

[0057]Corynebacterium acetoacidophilum ATCC13870

[0058]Corynebacterium thermoaminogenes FERM BP-1539

[0059]Corynebacterium melassecola ATCC17965

[0060]Brevibacterium flavum ATCC14067

[0061]Brevibacterium lactofermentum ATCC13869 and

[0062]Brevibacterium divaricatum ATCC14020

[0063] and L-amino acid-producing mutants or strains prepared therefrom, such as, for example, the L-lysine-producing strains

[0064]Corynebacterium glutamicum FERM-P 1709

[0065]Brevibacterium flavum FERM-P 1708

[0066]Brevibacterium lactofermentum FERM-P 1712

[0067]Corynebacterium glutamicum FERM-P 6463

[0068]Corynebacterium glutamicum FERM-P 6464

[0069]Corynebacterium glutamicum DM58-1

[0070]Corynebacterium glutamicum DG52-5

[0071]Corynebacterium glutamicum DSM5714 and

[0072]Corynebacterium glutamicum DSM12866.

[0073] Preferably, a bacterial strain with attenuated expression of a rpoB gene that encodes a polypeptide with activity of the β-subunit of RNA polymerase B will improve amino acid yield at least 1%.

[0074] The inventors have succeeded in isolating the new rpoB gene of C. glutamicum which codes for the β-subunit of RNA polymerase B, which is a β-subunit of RNA polymerase B.

[0075] In order to isolate the rpoB gene or other genes from C. glutamicum, a gene library of that microorganism in Escherichia coli (E. coli) is first prepared. The preparation of gene libraries is written down in generally known textbooks and handbooks. There may be mentioned as an example the textbook of Winnacker: Gene und Klone, Eine Einführung in die Gentechnologie (Verlag Chemie, Weinheim, Germany, 1990) or the handbook of Sambrook et al.: Molecular Cloning, A Laboratory Manual (Cold Spring Harbor Laboratory Press, 1989). A very well known gene library is that of the E. coli K-12 strain W3110, which has been prepared by Kohara et al. (Cell 50, 495-508 (1987)) in λ-vectors. Bathe et al. (Molecular and General Genetics, 252:255-265, 1996) describe a gene library of C. glutamicum ATCC13032, which has been prepared with the aid of the cosmid vector SuperCos I (Wahl et al., 1987, Proceedings of the National Academy of Sciences USA, 84:2160-2164) in the E. coli K-12 strain NM554 (Raleigh et al., 1988, Nucleic Acids Research 16:1563-1575).

[0076] Börmann et al. (Molecular Microbiology 6(3), 317-326) (sic) (1992)) in turn describe a gene library of C. glutamicum ATCC13032 using the cosmid pHC79 (Hohn and Collins, Gene 11, 291-298 (1980)).

[0077] For the preparation of a gene library of C. glutamicum in E. coli it is also possible to use plasmids such as pBR322 (Bolivar, Life Sciences, 25, 807-818 (1979)) or pUC9 (Vieira et al., 1982, Gene, 19:259-268). Suitable hosts are especially those E. coli strains that are restriction- and recombination-defective. An example thereof is the strain DH5αmcr, which has been described by Grant et al. (Proceedings of the National Academy of Sciences USA, 87 (1990) 4645-4649). The long DNA fragments cloned with the aid of cosmids can then in turn be subcloned into customary vectors suitable for sequencing and then sequenced, as is described, for example, in Sanger et al. (Proceedings of the National Academy of Sciences of the United States of America, 74:5463-5467, 1977).

[0078] The resulting DNA sequences can then be studied using known algorithms or sequence-analysis programs, such as, for example, that of Staden (Nucleic Acids Research 14, 217-232 (1986)), that of Marck (Nucleic Acids Research 16, 1829-1836 (1988)) or the GCG program of Butler (Methods of Biochemical Analysis 39, 74-97 (1998)).

[0079] The novel DNA sequence of C. glutamicum coding for the rpoB gene has been found and, as SEQ ID No. 1, forms part of the present invention. Furthermore, the amino acid sequence of the corresponding protein has been derived from the present DNA sequence using the methods described above. The resulting amino acid sequence of the rpoB gene product is shown in SEQ ID No. 2. It is known that enzymes belonging to the host are able to cleave the N-terminal amino acid methionine or formylmethionine of the protein that is formed.

[0080] Coding DNA sequences that result from SEQ ID No. 1 by the degeneracy of the genetic code also form part of the invention. Likewise, DNA sequences that hybridize with SEQ ID No. 1 or parts of SEQ ID No. 1 form part of the invention. Furthermore, to those skilled in the art, conservative amino acid substitutions, such as, for example, the substitution of glycine with alanine or of aspartic acid with glutamic acid, in proteins are known as sense mutations, which do not lead to any fundamental change in the activity of the protein, that is to say are neutral in terms of function. Such mutations are known inter alia also as neutral substitutions. It is also known that changes at the N- and/or C-terminus of a protein do not substantially impair its function or may even stabilise it. The person skilled in the art will find relevant information inter alia in Ben-Bassat et al. (Journal of Bacteriology 169:751-757 (1987)), in O'Regan et al. (Gene 77:237-251 (1989)), in Sahin-Toth et al. (Protein Sciences 3:240-247 (1994)), in Hochuli et al. (Bio/Technology 6:1321-1325 (1988)) and in known textbooks of genetics and molecular biology. Amino acid sequences that result in a corresponding manner from SEQ ID No. 2 likewise form part of the invention.

[0081] Similarly, DNA sequences that hybridize with SEQ ID No. 1 or parts of SEQ ID No. 1 form part of the invention. Finally, DNA sequences that are produced by the polymerase chain reaction (PCR) using primers that result from SEQ ID No. 1 form part of the invention. Such oligonucleotides typically have a length of at least 15 nucleotides.

[0082] The person skilled in the art will find instructions on the identification of DNA sequences by means of hybridization inter alia in the handbook “The DIG System Users Guide for Filter Hybridization” from Boehringer Mannheim GmbH (Mannheim, Germany, 1993) and in Liebl et al. (International Journal of Systematic Bacteriology (1991) 41: 255-260). The hybridization takes place under stringent conditions, that is to say there are formed only hybrids in which the probe and the target sequence, i.e. the polynucleotides treated with the probe, are at least 70% identical. It is known that the stringency of the hybridization, including the washing steps, is influenced or determined by varying the buffer composition, the temperature and the salt concentration. The hybridization reaction is preferably carried out with relatively low stringency as compared with the washing steps (Hybaid Hybridisation Guide, Hybaid Limited, Teddington, UK, 1996).

[0083] There may be used for the hybridization reaction, for example, a 5×SSC buffer at a temperature of approximately from 50° C. to 68° C. In that case, probes may also hybridize with polynucleotides that are less than 70% identical with the sequence of the probe. Such hybrids are less stable and are removed by washing under stringent conditions. That may be achieved, for example, by lowering the salt concentration to 2×SSC and optionally subsequently to 0.5×SSC (The DIG System User's Guide for Filter Hybridisation, Boehringer Mannheim, Mannheim, Germany, 1995), a temperature of approximately from 50° C. to 68° C. being set. It is optionally possible to lower the salt concentration down to 0.1×SSC. By raising the hybridization temperature stepwise from 50° C. to 68° C. in steps of approximately from 1 to 2° C., it is possible to isolate polynucleotide fragments that are, for example, at least 70% or at least 80% or at least from 90% to 95% identical with the sequence of the probe used. Further instructions for hybridization are commercially available in the form of so-called kits (e.g. DIG Easy Hyb from Roche Diagnostics GmbH, Mannheim, Germany, Catalog No. 1603558).

[0084] The person skilled in the art will find instructions on the amplification of DNA sequences with the aid of the polymerase chain reaction (PCR) inter alia in the handbook of Gait: Oligonukleotide synthesis: A Practical Approach (IRL Press, Oxford, UK, 1984) and in Newton and Graham: PCR (Spektrum Akademischer Verlag, Heidelberg, Germany, 1994).

[0085] It has been found that coryneform bacteria produce amino acids in an improved manner after enhancement of the rpoB gene.

[0086] In order to achieve overexpression, the number of copies of the corresponding genes can be increased, or the promoter and regulation region or the ribosome binding site, which is located upstream of the structural gene, can be mutated. Expression cassettes inserted upstream of the structural gene have a similar effect. By means of inducible promoters it is additionally possible to increase the expression in the course of the production of amino acids by fermentation. Expression is also improved by measures to prolong the life of the m-RNA. Furthermore, the enzyme activity is also enhanced by preventing degradation of the enzyme protein. The genes or gene constructs may either be present in plasmids with different numbers of copies or be integrated and amplified in the chromosome. Alternatively, overexpression of the genes in question may also be achieved by changing the composition of the medium and the manner in which culturing is carried out.

[0087] The person skilled in the art will find instructions thereon in Martin et al. (Bio/Technology 5, 137-146 (1987)), in Guerrero et al. (Gene 138, 35-41 (1994)), Tsuchiya and Morinaga (Bio/Technology 6, 428-430 (1988)), in Eikmanns et al. (Gene 102, 93-98 (1991)), in European patent specification 0 472 869, in U.S. Pat. No. 4,601,893, in Schwarzer and Pühler (Bio/Technology 9, 84-87 (1991), in Reinscheid et al. (Applied and Environmental Microbiology 60, 126-132 (1994)), in LaBarre et al. (Journal of Bacteriology 175, 1001-1007 (1993)), in patent application WO 96/15246, in Malumbres et al. (Gene 134, 15-24 (1993)), in Japanese Offenlegungsschrift JP-A-10-229891, in Jensen and Hammer (Biotechnology and Bioengineering 58, 191-195 (1998)), in Makrides (Microbiological Reviews 60:512-538 (1996)) and in known textbooks of genetics and molecular biology.

[0088] For the purposes of enhancement, the rpoB gene of the invention was overexpressed, for example, with the aid of episomal plasmids. Suitable plasmids are those which are replicated in coryneform bacteria. Many known plasmid vectors, such as, for example, pZ1 (Menkel et al., Applied and Environmental Microbiology (1989) 64: 549-554), pEKEx1 (Eikmanns et al., Gene 102:93-98 (1991)) or pHS2-1 (Sonnen et al., Gene 107:69-74 (1991)), are based on the cryptic plasmids pHM1519, pBL1 or pGA1. Other plasmid vectors, such as, for example, those which are based on pCG4 (U.S. Pat. No. 4,489,160) or pNG2 (Serwold-Davis et al., FEMS Microbiology Letters 66, 119-124 (1990)) or pAG1 (U.S. Pat. No. 5,158,891), may likewise be used.

[0089] Also suitable are those plasmid vectors with the aid of which the process of gene amplification by integration into the chromosome can be applied, as has been described, for example, by Reinscheid et al. (Applied and Environmental Microbiology 60, 126-132 (1994)) for the duplication or amplification of the hom-thrB operon. In that method, the complete gene is cloned into a plasmid vector that is able to replicate in a host (typically E. coli), but not in C. glutamicum. Suitable vectors are, for example, pSUP301 (Simon et al., Bio/Technology 1, 784-791 (1983)), pK18mob or pK19mob (Schäfer et al., Gene 145, 69-73 (1994)), pGEM-T (Promega corporation, Madison, Wis., USA), pCR2.1-TOPO (Shuman (1994). Journal of Biological Chemistry 269:32678-32684; U.S. Pat. No. 5,487,993), pCR®Blunt (Invitrogen, Groningen, Netherlands; Bernard et al., Journal of Molecular Biology, 234: 534-541 (1993)), pEM1 (Schrumpf et al., 1991, Journal of Bacteriology 173:4510-4516) or pBGS8 (Spratt et al., 1986, Gene 41: 337-342). The plasmid vector containing the gene to be amplified is then transferred to the desired strain of C. glutamicum by conjugation or transformation. The method of conjugation is described, for example, in Schäfer et al. (Applied and Environmental Microbiology 60, 756-759 (1994)). Methods of transformation are described, for example, in Thierbach et al. (Applied Microbiology and Biotechnology 29, 356-362 (1988)), Dunican and Shivnan (Bio/Technology 7, 1067-1070 (1989)) and Tauch et al. (FEMS Microbiological Letters 123, 343-347 (1994)). After homologous recombination by means of a “cross-over” occurrence, the resulting strain contains at least two copies of the gene in question.

[0090] It has also been found that the substitution of amino acids, especially in the sections between position 1 to 10, 190 to 200 and 420 to 450 in the amino acid sequence of the β-subunit of RNA polymerase B shown in SEQ ID No. 2, improves the lysine production of coryneform bacteria.

[0091] It has also been found that the substitution of amino acids at one or more positions selected from the group a) position 1 to 10, b) position 190 to 200 and c) position 420 to 450 in SEQ ID No. 2 may take place simultaneously.

[0092] In the region between position 1 to 10, preference is given to the substitution of L-proline at position 5 by L-leucine, L-isoleucine or L-valine.

[0093] In the region between position 190 to 200, preference is given to the substitution of L-serine at position 196 by L-phenylalanine or L-tyrosine.

[0094] In the region between 420 to 450, the following substitutions are preferred: substitution of L-leucine at position 424 by L-proline or L-arginine, substitution of L-serine at position 425 by L-threonine or L-alanine, substitution of L-glutamine at position 426 by L-leucine or L-lysine, substitution of L-aspartic acid at position 429 by L-isoleucine, L-valine or L-leucine, substitution of L-histidine at position 439 by any proteinogenic amino acid with the exception of L-histidine, is (sic) the substitution of L-serine at position 444 by L-leucine, L-tyrosine or L-tryptophan, and substitution of L-leucine at position 446 by L-proline or L-isoleucine.

[0095] Very special preference is given to one or more amino acid substitutions selected from the group: L-proline at position 5 by L-leucine, L-serine at position 196 by L-phenylalanine, L-aspartate at position 429 by L-valine, and L-histidine at position 439 by L-tyrosine.

[0096] SEQ ID No. 3 shows the base sequence of the allele rpoB-1547 contained in strain DM1547. The rpoB-1547 allele codes for a protein the amino acid sequence of which is shown in SEQ ID No. 4. The protein contains L-leucine at position 5, L-phenylalanine at position 196 and L-valine at position 429. The DNA sequence of the rpoB-1547 allele (SEQ ID No. 3) contains the following base substitutions as compared with the rpoB wild-type gene (SEQ ID No. 1): thymine at position 715 instead of cytosine, thymine at position 1288 instead of cytosine, and thymine at position 1987 instead of adenine.

[0097] SEQ ID No. 5 shows the base sequence of the allele rpoB-1546 contained in strain DM1546. The rpoB-1546 allele codes for a protein the amino acid sequence of which is shown in SEQ ID No. 6. The protein contains L-tyrosine at position 439. The DNA sequence of the rpoB-1546 allele (SEQ ID No. 5) contains the following base substitutions as compared with the rpoB wild-type gene (SEQ ID No. 1): thymine at position 2016 instead of cytosine.

[0098] There may be employed for the mutagenesis conventional methods of mutagenesis using mutagenic substances such as, for example, N-methyl-N′-nitro-N-nitrosoguanidine or ultraviolet light. There may also be used for the mutagenesis in vitro methods such as, for example, treatment with hydroxylamine (Miller, J. H.: A Short Course in Bacterial Genetics. A Laboratory Manual and Handbook for Escherichia coli and Related Bacteria, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, 1992) or mutagenic oligonucleotides (T. A. Brown: Gentechnologie für Einsteiger, Spektrum Akademischer Verlag, Heidelberg, 1993) or the polymerase chain reaction (PCR), as is described in the handbook of Newton and Graham (PCR, Spektrum Akademischer Verlag, Heidelberg, 1994).

[0099] In addition, it may be advantageous for the production of L-amino acids to enhance, especially to overexpress, in addition to the rpoB gene, one or more enzymes of the biosynthesis pathway in question, of glycolysis, of the anaplerotic pathway, of the citric acid cycle, of the pentose phosphate cycle, of amino acid export, and, optionally, regulatory proteins.

[0100] Accordingly, for the production of L-lysine, in addition to enhancing the rpoB gene, one or more genes selected from the group

[0101] the gene dapA coding for dihydrodipicolinate synthase (EP-B 0 197 335),

[0102] the gene gap coding for glyceraldehyde 3-phosphate dehydrogenase (Eikmanns (1992), Journal of Bacteriology 174:6076-6086),

[0103] the gene tpi coding for triose phosphate isomerase (Eikmanns (1992), Journal of Bacteriology 174:6076-6086),

[0104] the gene pgk coding for 3-phosphoglycerate kinase (Eikmanns (1992), Journal of Bacteriology 174:6076-6086),

[0105] the gene zwf coding for glucose-6-phosphate dehydrogenase (JP-A-09224661),

[0106] the gene pyc coding for pyruvate carboxylase (DE-A-198 31 609),

[0107] the gene mqo coding for malate quinone oxidoreductase (Molenaar et al., European Journal of Biochemistry 254, 395-403 (1998)),

[0108] the gene lysC coding for a feed-back resistant aspartate kinase (Kalinowski et al., Molecular Microbiologie 5(5), 1197-1204 (1991)),

[0109] the gene lysE coding for lysine export (DE-A-195 48 222),

[0110] the gene zwa1 coding for the Zwa1 protein (DE: 19959328.0, DSM 13115), and

[0111] the rpsL gene coding for ribosomal protein S12 and shown in SEQ ID No. 7 and 8

[0112] may be enhanced, especially overexpressed.

[0113] The term “attenuation” in this connection describes the diminution or exclusion of the intracellular activity of one or more enzymes (proteins) in a microorganism that are coded for by the corresponding DNA, by, for example, using a weak promoter or using a gene or allele that codes for a corresponding enzyme having low activity, or by inactivating the corresponding gene or enzyme (protein), and optionally by combining those measures.

[0114] Furthermore, it may be advantageous for the production of L-amino acids, in addition to enhancing the rpob gene, to attenuate, especially to diminish the expression of, one or more genes selected from the group

[0115] the gene pck coding for phosphoenol pyruvate carboxykinase (DE 199 50 409.1; DSM 13047),

[0116] the gene pgi coding for glucose-6-phosphate isomerase (U.S. 09/396,478; DSM 12969),

[0117] the gene poxB coding for pyruvate oxidase (DE: 1995 1975.7; DSM 13114),

[0118] the gene zwa2 coding for the Zwa2 protein (DE: 19959327.2, DSM 13113).

[0119] It may also be advantageous for the production of amino acids, in addition to enhancing the rpoB gene, to exclude undesired secondary reactions (Nakayama: “Breeding of Amino Acid Producing Micro-organisms”, in: Overproduction of Microbial Products, Krumphanzl, Sikyta, Vanek (eds.), Academic Press, London, UK, 1982).

[0120] The microorganisms produced according to the invention also form part of the invention and can be cultivated, for the purposes of the production of amino acids, continuously or discontinuously in the batch, fed batch or repeated fed batch process. A summary of known cultivation methods is described in the textbook of Chmiel (Bioprozeβtechnik 1. Einführung in die Bioverfahrenstechnik (Gustav Fischer Verlag, Stuttgart, 1991)) or in the textbook of Storhas (Bioreaktoren und periphere Einrichtungen (Vieweg Verlag, Braunschweig/Wiesbaden, 1994)).

[0121] The culture medium to be used must meet the requirements of the strains in question in a suitable manner. Descriptions of culture media for various microorganisms are to be found in the handbook “Manual of Methods for General Bacteriology” of the American Society for Bacteriology (Washington D.C., USA, 1981).

[0122] There may be used as the carbon source sugars and carbohydrates, such as, for example, glucose, saccharose, lactose, fructose, maltose, molasses, starch and cellulose, oils and fats, such as, for example, soybean oil, sunflower oil, groundnut oil and coconut oil, fatty acids, such as, for example, palmitic acid, stearic acid and linoleic acid, alcohols, such as, for example, glycerol and ethanol, and organic acids, such as, for example, acetic acid. Those substances may be used individually or in the form of a mixture.

[0123] There may be used as the nitrogen source organic nitrogen-containing compounds, such as peptones, yeast extract, meat extract, malt extract, corn steep liquor, soybean flour and urea, or inorganic compounds, such as ammonium sulfate, ammonium chloride, ammonium phosphate, ammonium carbonate and ammonium nitrate. The nitrogen sources may be used individually or in the form of a mixture.

[0124] There may be used as the phosphorus source phosphoric acid, potassium dihydrogen phosphate or dipotassium hydrogen phosphate or the corresponding sodium-containing salts. The culture medium must also contain salts of metals, such as, for example, magnesium sulfate or iron sulfate, which are necessary for growth. Finally, essential growth substances, such as amino acids and vitamins, may be used in addition to the above-mentioned substances. Suitable precursors may also be added to the culture medium. The mentioned substances may be added to the culture in the form of a single batch, or they may be fed in in a suitable manner during the cultivation.

[0125] In order to control the pH value of the culture, basic compounds, such as sodium hydroxide, potassium hydroxide, ammonia or ammonia water, or acid compounds, such as phosphoric acid or sulfuric acid, are expediently used. In order to control the development of foam, anti-foams, such as, for example, fatty acid polyglycol esters, may be used. In order to maintain the stability of plasmids, suitable substances having a selective action, such as, for example, antibiotics, may be added to the medium. In order to maintain aerobic conditions, oxygen or gas mixtures containing oxygen, such as, for example, air, are introduced into the culture. The temperature of the culture is normally from 20° C. to 45° C. and preferably from 25° C. to 40° C. The culture is continued until the maximum amount of the desired product has formed. That aim is normally achieved within a period of from 10 hours to 160 hours.

[0126] Methods of determining L-amino acids are known from the prior art. The analysis may be carried out, for example, as described in Spackman et al. (Analytical Chemistry, 30, (1958), 1190) by ion-exchange chromatography with subsequent ninhydrin derivatization, or it may be carried out by reversed phase HPLC, as described in Lindroth et al. (Analytical Chemistry (1979) 51: 1167-1174).

[0127] Pure cultures of the following microorganisms were deposited on 16 Jan. 2001 at the Deutsche Sammlung für Mikroorganismen und Zellkulturen (DSMZ, Braunschweig, Germany) in accordance with the Budapest Treaty:

[0128]Corynebacterium glutamicum strain DM1546 as DSM 13993

[0129]Corynebacterium glutamicum strain DM1547 as DSM 13994.

[0130] The process of the invention is used for the production of amino acids by fermentation.

[0131] The present invention is explained in greater detail below by means of Examples.

[0132] The isolation of plasmid DNA from Escherichia coli and all techniques for restriction, Klenow and alkaline phosphatase treatment were carried out according to Sambrook et al. (Molecular Cloning. A Laboratory Manual (1989) Cold Spring Harbour Laboratory Press, Cold Spring Harbor, N.Y., USA). Methods for the transformation of Escherichia coli are also described in that handbook.

[0133] The composition of common nutrient media, such as LB or TY medium, will also be found in the handbook of Sambrook et al.

EXAMPLE 1

[0134] Preparation of a genomic cosmid gene library from Corynebacterium glutamicum ATCC 13032

[0135] Chromosomal DNA from Corynebacterium glutamicum ATCC 13032 is isolated as described in Tauch et al. (1995, Plasmid 33:168-179) and partially cleaved with the restriction enzyme Sau3AI (Amersham Pharmacia, Freiburg, Germany, product description Sau3AI, Code no. 27-0913-02). The DNA fragments are dephosphorylated with shrimp alkaline phosphatase (Roche Diagnostics GmbH, Mannheim, Germany, product description SAP, Code no. 1758250). The DNA of cosmid vector SuperCos1 (Wahl et al. (1987) Proceedings of the National Academy of Sciences USA 84:2160-2164), obtained from Stratagene (La Jolla, USA, product description SuperCos1 Cosmid Vektor Kit, Code no. 251301), is cleaved with the restriction enzyme XbaI (Amersham Pharmacia, Freiburg, Germany, product description XbaI, Code no. 27-0948-02) and likewise dephosphorylated with shrimp alkaline phosphatase.

[0136] The cosmid DNA is then cleaved with the restriction enzyme BamHI (Amersham Pharmacia, Freiburg, Germany, product description BamHI, Code no. 27-0868-04). The cosmid DNA so treated is mixed with the treated ATCC13032 DNA, and the batch is treated with T4-DNA ligase (Amersham Pharmacia, Freiburg, Germany, product description T4-DNA ligase, Code no. 27-0870-04). The ligation mixture is then packed in phages with the aid of Gigapack II XL Packing Extract (Stratagene, La Jolla, USA, product description Gigapack II XL Packing Extract, Code no. 200217).

[0137] For infection of E. coli strain NM554 (Raleigh et al. 1988, Nucleic Acid Research 16:1563-1575), the cells are taken up in 10 mM MgSO₄ and mixed with an aliquot of the phage suspension. Infection and titration of the cosmid library are carried out as described in Sambrook et al. (1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor), the cells being plated out on LB agar (Lennox, 1955, Virology, 1:190) with 100 mg/l ampicillin. After incubation overnight at 37° C., recombinant individual clones are selected.

EXAMPLE 2

[0138] Isolation and sequencing of the rpoB gene

[0139] The cosmid DNA of an individual colony is isolated using the Qiaprep Spin Miniprep Kit (Product No. 27106, Qiagen, Hilden, Germany) according to the. manufacturer's instructions, and partially cleaved with the restriction enzyme Sau3AI (Amersham Pharmacia, Freiburg, Germany, product description Sau3AI, Product No. 27-0913-02). The DNA fragments are dephosphorylated with shrimp alkaline phosphatase (Roche Diagnostics GmbH, Mannheim, Germany, product description SAP, Product No. 1758250). After separation by gel electrophoresis, cosmid fragments having a size in the range from 1500 to 2000 bp are isolated using the QiaExII Gel Extraction Kit (Product No. 20021, Qiagen, Hilden, Germany).

[0140] The DNA of sequencing vector pZero-1, obtained from Invitrogen (Groningen, Netherlands, product description Zero Background Cloning Kit, Product No. K2500-01), is cleaved with the restriction enzyme BamHI (Amersham Pharmacia, Freiburg, Germany, product description BamHI, Product No. 27-0868-04). Ligation of the cosmid fragments into the sequencing vector pzero-1 is carried out as described by Sambrook et al. (1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor), the DNA mixture being incubated overnight with T4 ligase (Pharmacia Biotech, Freiburg, Germany). The ligation mixture is then electroporated into E. coli strain DH5αMCR (Grant, 1990, Proceedings of the National Academy of Sciences U.S.A., 87:4645-4649) (Tauch et al. 1994, FEMS Microbiol Letters, 123:343-347) and plated out on LB agar (Lennox, 1955, Virology, 1:190) with 50 mg/l Zeocin.

[0141] Plasmid preparation of the recombinant clones is carried out using the Biorobot 9600 (Product No. 900200, Qiagen, Hilden, Germany). Sequencing is effected by the dideoxy chain termination method of Sanger et al. (1977, Proceedings of the National Academy of Sciences U.S.A., 74:5463-5467) with modifications according to Zimmermann et al. (1990, Nucleic Acids Research, 18:1067). The “RR dRhodamin Terminator Cycle Sequencing Kit” from PE Applied Biosystems (Product No. 403044, Weiterstadt, Germany) is used. Separation by gel electrophoresis and analysis of the sequencing reaction is carried out in a “Rotiphorese NF Acrylamid/Bisacrylamid” gel (29:1) (Product No. A124.1, Roth, Karlsruhe, Germany) using the “ABI Prism 377” sequencing device from PE Applied Biosystems (Weiterstadt, Germany).

[0142] The resulting crude sequence data are then processed using the Staden program package (1986, Nucleic Acids Research, 14:217-231) Version 97-0. The individual sequences of the pzerol derivatives are assembled to a coherent contig. The computer-assisted coding region analysis is prepared using the program XNIP (Staden, 1986, Nucleic Acids Research, 14:217-231)

[0143] The resulting nucleotide sequence is shown in SEQ ID No. 1. Analysis of the nucleotide sequence gives an open reading frame of 3497 base pairs, which is designated the rpoB gene. The rpoB gene codes for a protein of 1165 amino acids.

[0144] Obviously, numerous modifications and variations on the present invention are possible in light of the above teachings. It is therefore to be understood that within the scope of the appended claims, the invention may be practiced otherwise than as specifically described herein.

1 12 1 5099 DNA Corynebacterium glutamicum CDS (702)..(4196) 1 acaatgtgac tcgtgatttt tgggtggatc agcgtaccgg tttggttgtc gatctagctg 60 aaaatattga tgatttttac ggcgaccgca gcggccagaa gtacgaacag aaattgcttt 120 tcgacgcctc cctcgacgat gcagctgtct ctaagctggt tgcacaggcc gaaagcatcc 180 ctgatggaga tgtgagcaaa atcgcaaata ccgtaggtat tgtgatcggt gcggtattgg 240 ctctcgtggg cctggccggg tgttttgggg cgtttgggaa gaaacgtcga gaagcttaac 300 ctgctgttca aatagatttt ccctgtttcg aattgcggaa accccgggtt tgtttgctag 360 ggtgcctcgt agaaggggtc aagaagattt ctgggaaacg cgcccgtgcg gttggttgct 420 aatagcacgc ggagcaccag atgaaaaatc tcccctttac tttcgcgcgc gattggtata 480 ctctgagtcg ttgcgttgga attcgtgact ctttttcgtt cctgtagcgc caagaccttg 540 atcaaggtgg tttaaaaaaa ccgatttgac aaggtcattc agtgctatct ggagtcgttc 600 agggggatcg ggttcctcag cagaccaatt gctcaaaaat accagcggtg ttgatctgca 660 cttaatggcc ttgaccagcc aggtgcaatt acccgcgtga g gtg ctg gaa gga ccc 716 Val Leu Glu Gly Pro 1 5 atc ttg gca gtc tcc cgc cag acc aag tca gtc gtc gat att ccc ggt 764 Ile Leu Ala Val Ser Arg Gln Thr Lys Ser Val Val Asp Ile Pro Gly 10 15 20 gca ccg cag cgt tat tct ttc gcg aag gtg tcc gca ccc att gag gtg 812 Ala Pro Gln Arg Tyr Ser Phe Ala Lys Val Ser Ala Pro Ile Glu Val 25 30 35 ccc ggg cta cta gat ctt caa ctg gat tct tac tcc tgg ctg att ggt 860 Pro Gly Leu Leu Asp Leu Gln Leu Asp Ser Tyr Ser Trp Leu Ile Gly 40 45 50 acg cct gag tgg cgt gct cgt cag aag gaa gaa ttc ggc gag gga gcc 908 Thr Pro Glu Trp Arg Ala Arg Gln Lys Glu Glu Phe Gly Glu Gly Ala 55 60 65 cgc gta acc agc ggc ctt gag aac att ctc gag gag ctc tcc cca atc 956 Arg Val Thr Ser Gly Leu Glu Asn Ile Leu Glu Glu Leu Ser Pro Ile 70 75 80 85 cag gat tac tct gga aac atg tcc ctg agc ctt tcg gag cca cgc ttc 1004 Gln Asp Tyr Ser Gly Asn Met Ser Leu Ser Leu Ser Glu Pro Arg Phe 90 95 100 gaa gac gtc aag aac acc att gac gag gcg aaa gaa aag gac atc aac 1052 Glu Asp Val Lys Asn Thr Ile Asp Glu Ala Lys Glu Lys Asp Ile Asn 105 110 115 tac gcg gcg cca ctg tat gtg acc gcg gag ttc gtc aac aac acc acc 1100 Tyr Ala Ala Pro Leu Tyr Val Thr Ala Glu Phe Val Asn Asn Thr Thr 120 125 130 ggt gaa atc aag tct cag act gtc ttc atc ggc gat ttc cca atg atg 1148 Gly Glu Ile Lys Ser Gln Thr Val Phe Ile Gly Asp Phe Pro Met Met 135 140 145 acg gac aag gga acg ttc atc atc aac gga acc gaa cgc gtt gtg gtc 1196 Thr Asp Lys Gly Thr Phe Ile Ile Asn Gly Thr Glu Arg Val Val Val 150 155 160 165 agc cag ctc gtc cgc tcc ccg ggc gtg tac ttt gac cag acc atc gat 1244 Ser Gln Leu Val Arg Ser Pro Gly Val Tyr Phe Asp Gln Thr Ile Asp 170 175 180 aag tca act gag cgt cca ctg cac gcc gtg aag gtt att cct tcc cgt 1292 Lys Ser Thr Glu Arg Pro Leu His Ala Val Lys Val Ile Pro Ser Arg 185 190 195 ggt gct tgg ctt gag ttt gac gtc gat aag cgc gat tcg gtt ggt gtt 1340 Gly Ala Trp Leu Glu Phe Asp Val Asp Lys Arg Asp Ser Val Gly Val 200 205 210 cgt att gac cgc aag cgt cgc cag cca gtc acc gta ctg ctg aag gct 1388 Arg Ile Asp Arg Lys Arg Arg Gln Pro Val Thr Val Leu Leu Lys Ala 215 220 225 ctt ggc tgg acc act gag cag atc acc gag cgt ttc ggt ttc tct gaa 1436 Leu Gly Trp Thr Thr Glu Gln Ile Thr Glu Arg Phe Gly Phe Ser Glu 230 235 240 245 atc atg atg tcc acc ctc gag tcc gat ggt gta gca aac acc gat gag 1484 Ile Met Met Ser Thr Leu Glu Ser Asp Gly Val Ala Asn Thr Asp Glu 250 255 260 gca ttg ctg gag atc tac cgc aag cag cgt cca ggc gag cag cct acc 1532 Ala Leu Leu Glu Ile Tyr Arg Lys Gln Arg Pro Gly Glu Gln Pro Thr 265 270 275 cgc gac ctt gcg cag tcc ctc ctg gac aac agc ttc ttc cgt gca aag 1580 Arg Asp Leu Ala Gln Ser Leu Leu Asp Asn Ser Phe Phe Arg Ala Lys 280 285 290 cgc tac gac ctg gct cgc gtt ggt cgt tac aag atc aac cgc aag ctc 1628 Arg Tyr Asp Leu Ala Arg Val Gly Arg Tyr Lys Ile Asn Arg Lys Leu 295 300 305 ggc ctt ggt ggc gac cac gat ggt ttg atg act ctt act gaa gag gac 1676 Gly Leu Gly Gly Asp His Asp Gly Leu Met Thr Leu Thr Glu Glu Asp 310 315 320 325 atc gca acc acc atc gag tac ctg gtg cgt ctg cac gca ggt gag cgc 1724 Ile Ala Thr Thr Ile Glu Tyr Leu Val Arg Leu His Ala Gly Glu Arg 330 335 340 gtc atg act tct cca aat ggt gaa gag atc cca gtc gag acc gat gac 1772 Val Met Thr Ser Pro Asn Gly Glu Glu Ile Pro Val Glu Thr Asp Asp 345 350 355 atc gac cac ttt ggt aac cgt cgt ctg cgt acc gtt ggc gaa ctg atc 1820 Ile Asp His Phe Gly Asn Arg Arg Leu Arg Thr Val Gly Glu Leu Ile 360 365 370 cag aac cag gtc cgt gtc ggc ctg tcc cgc atg gag cgc gtt gtt cgt 1868 Gln Asn Gln Val Arg Val Gly Leu Ser Arg Met Glu Arg Val Val Arg 375 380 385 gag cgt atg acc acc cag gat gcg gag tcc att act cct act tcc ttg 1916 Glu Arg Met Thr Thr Gln Asp Ala Glu Ser Ile Thr Pro Thr Ser Leu 390 395 400 405 atc aac gtt cgt cct gtc tct gca gct atc cgt gag ttc ttc gga act 1964 Ile Asn Val Arg Pro Val Ser Ala Ala Ile Arg Glu Phe Phe Gly Thr 410 415 420 tcc cag ctg tct cag ttc atg gac cag aac aac tcc ctg tct ggt ttg 2012 Ser Gln Leu Ser Gln Phe Met Asp Gln Asn Asn Ser Leu Ser Gly Leu 425 430 435 act cac aag cgt cgt ctg tcg gct ctg ggc ccg ggt ggt ctg tcc cgt 2060 Thr His Lys Arg Arg Leu Ser Ala Leu Gly Pro Gly Gly Leu Ser Arg 440 445 450 gag cgc gcc ggc atc gag gtt cga gac gtt cac cca tct cac tac ggc 2108 Glu Arg Ala Gly Ile Glu Val Arg Asp Val His Pro Ser His Tyr Gly 455 460 465 cgt atg tgc cca att gag act ccg gaa ggt cca aac att ggc ctg atc 2156 Arg Met Cys Pro Ile Glu Thr Pro Glu Gly Pro Asn Ile Gly Leu Ile 470 475 480 485 ggt tcc ttg gct tcc tat gct cga gtg aac cca ttc ggt ttc att gag 2204 Gly Ser Leu Ala Ser Tyr Ala Arg Val Asn Pro Phe Gly Phe Ile Glu 490 495 500 acc cca tac cgt cgc atc atc gac ggc aag ctg acc gac cag att gac 2252 Thr Pro Tyr Arg Arg Ile Ile Asp Gly Lys Leu Thr Asp Gln Ile Asp 505 510 515 tac ctt acc gct gat gag gaa gac cgc ttc gtt gtt gcg cag gca aac 2300 Tyr Leu Thr Ala Asp Glu Glu Asp Arg Phe Val Val Ala Gln Ala Asn 520 525 530 acg cac tac gac gaa gag ggc aac atc acc gat gag acc gtc act gtt 2348 Thr His Tyr Asp Glu Glu Gly Asn Ile Thr Asp Glu Thr Val Thr Val 535 540 545 cgt ctg aag gac ggc gac atc gcc atg gtt ggc cgc aac gcg gtt gat 2396 Arg Leu Lys Asp Gly Asp Ile Ala Met Val Gly Arg Asn Ala Val Asp 550 555 560 565 tac atg gac gtt tcc cct cgt cag atg gtt tct gtt ggt acc gcg atg 2444 Tyr Met Asp Val Ser Pro Arg Gln Met Val Ser Val Gly Thr Ala Met 570 575 580 att cca ttc ctg gag cac gac gat gct aac cgt gca ctg atg ggc gcg 2492 Ile Pro Phe Leu Glu His Asp Asp Ala Asn Arg Ala Leu Met Gly Ala 585 590 595 aac atg cag aag cag gct gtg cca ctg att cgt gcc gag gct cct ttc 2540 Asn Met Gln Lys Gln Ala Val Pro Leu Ile Arg Ala Glu Ala Pro Phe 600 605 610 gtg ggc acc ggt atg gag cag cgc gca gca tac gac gcc ggc gac ctg 2588 Val Gly Thr Gly Met Glu Gln Arg Ala Ala Tyr Asp Ala Gly Asp Leu 615 620 625 gtt att acc cca gtc gca ggt gtg gtg gaa aac gtt tca gct gac ttc 2636 Val Ile Thr Pro Val Ala Gly Val Val Glu Asn Val Ser Ala Asp Phe 630 635 640 645 atc acc atc atg gct gat gac ggc aag cgc gaa acc tac ctg ctg cgt 2684 Ile Thr Ile Met Ala Asp Asp Gly Lys Arg Glu Thr Tyr Leu Leu Arg 650 655 660 aag ttc cag cgc acc aac cag ggc acc agc tac aac cag aag cct ttg 2732 Lys Phe Gln Arg Thr Asn Gln Gly Thr Ser Tyr Asn Gln Lys Pro Leu 665 670 675 gtt aac ttg ggc gag cgc gtt gaa gct ggc cag gtt att gct gat ggt 2780 Val Asn Leu Gly Glu Arg Val Glu Ala Gly Gln Val Ile Ala Asp Gly 680 685 690 cca ggt acc ttc aat ggt gaa atg tcc ctt ggc cgt aac ctt ctg gtt 2828 Pro Gly Thr Phe Asn Gly Glu Met Ser Leu Gly Arg Asn Leu Leu Val 695 700 705 gcg ttc atg cct tgg gaa ggc cac aac tac gag gat gcg atc atc ctc 2876 Ala Phe Met Pro Trp Glu Gly His Asn Tyr Glu Asp Ala Ile Ile Leu 710 715 720 725 aac cag aac atc gtt gag cag gac atc ttg acc tcg atc cac atc gag 2924 Asn Gln Asn Ile Val Glu Gln Asp Ile Leu Thr Ser Ile His Ile Glu 730 735 740 gag cac gag atc gat gcc cgc gac act aag ctt ggc gcc gaa gaa atc 2972 Glu His Glu Ile Asp Ala Arg Asp Thr Lys Leu Gly Ala Glu Glu Ile 745 750 755 acc cgc gac atc cct aat gtg tct gaa gaa gtc ctc aag gac ctc gac 3020 Thr Arg Asp Ile Pro Asn Val Ser Glu Glu Val Leu Lys Asp Leu Asp 760 765 770 gac cgc ggt att gtc cgc atc ggt gct gat gtt cgt gac ggc gac atc 3068 Asp Arg Gly Ile Val Arg Ile Gly Ala Asp Val Arg Asp Gly Asp Ile 775 780 785 ctg gtc ggt aag gtc acc cct aag ggc gag acc gag ctc acc ccg gaa 3116 Leu Val Gly Lys Val Thr Pro Lys Gly Glu Thr Glu Leu Thr Pro Glu 790 795 800 805 gag cgc ttg ctg cgc gca atc ttc ggt gag aag gcc cgc gaa gtt cgc 3164 Glu Arg Leu Leu Arg Ala Ile Phe Gly Glu Lys Ala Arg Glu Val Arg 810 815 820 gat acc tcc atg aag gtg cct cac ggt gag acc ggc aag gtc atc ggc 3212 Asp Thr Ser Met Lys Val Pro His Gly Glu Thr Gly Lys Val Ile Gly 825 830 835 gtg cgt cac ttc tcc cgc gag gac gac gac gat ctg gct cct ggc gtc 3260 Val Arg His Phe Ser Arg Glu Asp Asp Asp Asp Leu Ala Pro Gly Val 840 845 850 aac gag atg atc cgt atc tac gtt gct cag aag cgt aag atc cag gac 3308 Asn Glu Met Ile Arg Ile Tyr Val Ala Gln Lys Arg Lys Ile Gln Asp 855 860 865 ggc gat aag ctc gct ggc cgc cac ggt aac aag ggt gtt gtc ggt aaa 3356 Gly Asp Lys Leu Ala Gly Arg His Gly Asn Lys Gly Val Val Gly Lys 870 875 880 885 att ttg cct cag gaa gat atg cca ttc ctt cca gac ggc act cct gtt 3404 Ile Leu Pro Gln Glu Asp Met Pro Phe Leu Pro Asp Gly Thr Pro Val 890 895 900 gac atc atc ttg aac acc cac ggt gtt cca cgt cgt atg aac att ggt 3452 Asp Ile Ile Leu Asn Thr His Gly Val Pro Arg Arg Met Asn Ile Gly 905 910 915 cag gtt ctt gag acc cac ctt ggc tgg ctg gca tct gct ggt tgg tcc 3500 Gln Val Leu Glu Thr His Leu Gly Trp Leu Ala Ser Ala Gly Trp Ser 920 925 930 gtg gat cct gaa gat cct gag aac gct gag ctc gtc aag act ctg cct 3548 Val Asp Pro Glu Asp Pro Glu Asn Ala Glu Leu Val Lys Thr Leu Pro 935 940 945 gca gac ctc ctc gag gtt cct gct ggt tcc ttg act gca act cct gtg 3596 Ala Asp Leu Leu Glu Val Pro Ala Gly Ser Leu Thr Ala Thr Pro Val 950 955 960 965 ttc gac ggt gcg tca aac gaa gag ctc gca ggc ctg ctc gct aat tca 3644 Phe Asp Gly Ala Ser Asn Glu Glu Leu Ala Gly Leu Leu Ala Asn Ser 970 975 980 cgt cca aac cgc gac ggc gac gtc atg gtt aac gcg gat ggt aaa gca 3692 Arg Pro Asn Arg Asp Gly Asp Val Met Val Asn Ala Asp Gly Lys Ala 985 990 995 acg ctt atc gac ggt cgc tcc ggt gag cct tac ccg tac ccg gtt 3737 Thr Leu Ile Asp Gly Arg Ser Gly Glu Pro Tyr Pro Tyr Pro Val 1000 1005 1010 tcc atc ggc tac atg tac atg ctg aag ctg cac cac ctc gtt gac 3782 Ser Ile Gly Tyr Met Tyr Met Leu Lys Leu His His Leu Val Asp 1015 1020 1025 gag aag atc cac gca cgt tcc act ggt cct tac tcc atg att acc 3827 Glu Lys Ile His Ala Arg Ser Thr Gly Pro Tyr Ser Met Ile Thr 1030 1035 1040 cag cag cca ctg ggt ggt aaa gca cag ttc ggt gga cag cgt ttc 3872 Gln Gln Pro Leu Gly Gly Lys Ala Gln Phe Gly Gly Gln Arg Phe 1045 1050 1055 ggc gaa atg gag gtg tgg gca atg cag gca tac ggc gct gcc tac 3917 Gly Glu Met Glu Val Trp Ala Met Gln Ala Tyr Gly Ala Ala Tyr 1060 1065 1070 aca ctt cag gag ctg ctg acc atc aag tct gat gac gtg gtt ggc 3962 Thr Leu Gln Glu Leu Leu Thr Ile Lys Ser Asp Asp Val Val Gly 1075 1080 1085 cgt gtc aag gtc tac gaa gca att gtg aag ggc gag aac atc ccg 4007 Arg Val Lys Val Tyr Glu Ala Ile Val Lys Gly Glu Asn Ile Pro 1090 1095 1100 gat cca ggt att cct gag tcc ttc aag gtt ctc ctc aag gag ctc 4052 Asp Pro Gly Ile Pro Glu Ser Phe Lys Val Leu Leu Lys Glu Leu 1105 1110 1115 cag tcc ttg tgc ctg aac gtg gag gtt ctc tcc gca gac ggc act 4097 Gln Ser Leu Cys Leu Asn Val Glu Val Leu Ser Ala Asp Gly Thr 1120 1125 1130 cca atg gag ctc gcg ggt gac gac gac gac ttc gat cag gca ggc 4142 Pro Met Glu Leu Ala Gly Asp Asp Asp Asp Phe Asp Gln Ala Gly 1135 1140 1145 gcc tca ctt ggc atc aac ctg tcc cgt gac gag cgt tcc gac gcc 4187 Ala Ser Leu Gly Ile Asn Leu Ser Arg Asp Glu Arg Ser Asp Ala 1150 1155 1160 gac acc gca tagcagatca gaaaacaacc gctagaaatc aagccataca 4236 Asp Thr Ala 1165 tcccccggac attgaagaga tgttctgggg ggaaagggag ttttacgtgc tcgacgtaaa 4296 cgtcttcgat gagctccgca tcggcctggc caccgccgac gacatccgcc gttggtccaa 4356 gggtgaggtc aagaagccgg agaccatcaa ctaccgaacc ctcaagcctg agaaggacgg 4416 tctgttctgc gagcgtatct tcggtccaac tcgcgactgg gagtgcgcct gcggtaagta 4476 caagcgtgtc cgctacaagg gcatcatctg tgaacgctgt ggcgttgagg tcaccaagtc 4536 caaggtgcgc cgtgagcgca tgggacacat tgagctcgct gcaccagtaa cccacatttg 4596 gtacttcaag ggcgttccat cacgcctcgg ctaccttttg gaccttgctc caaaggacct 4656 ggacctcatc atctacttcg gtgcgaacat catcaccagc gtggacgaag aggctcgcca 4716 cagcgaccag accactcttg aggcagaaat gcttctggag aagaaggacg ttgaggcaga 4776 cgcagagtct gacattgctg agcgtgctga aaagctcgaa gaggatcttg ctgaacttga 4836 ggcagctggc gctaaggccg acgctcgccg caaggttcag gctgctgccg ataaggaaat 4896 gcagcacatc cgtgagcgtg cacagcgcga aatcgatcgt ctcgatgagg tctggcagac 4956 cttcatcaag cttgctccaa agcagatgat ccgcgatgag aagctctacg atgaactgat 5016 cgaccgctac gaggattact tcaccggtgg tatgggtgca gagtccattg aggctttgat 5076 ccagaacttc gaccttgatg ctg 5099 2 1165 PRT Corynebacterium glutamicum 2 Val Leu Glu Gly Pro Ile Leu Ala Val Ser Arg Gln Thr Lys Ser Val 1 5 10 15 Val Asp Ile Pro Gly Ala Pro Gln Arg Tyr Ser Phe Ala Lys Val Ser 20 25 30 Ala Pro Ile Glu Val Pro Gly Leu Leu Asp Leu Gln Leu Asp Ser Tyr 35 40 45 Ser Trp Leu Ile Gly Thr Pro Glu Trp Arg Ala Arg Gln Lys Glu Glu 50 55 60 Phe Gly Glu Gly Ala Arg Val Thr Ser Gly Leu Glu Asn Ile Leu Glu 65 70 75 80 Glu Leu Ser Pro Ile Gln Asp Tyr Ser Gly Asn Met Ser Leu Ser Leu 85 90 95 Ser Glu Pro Arg Phe Glu Asp Val Lys Asn Thr Ile Asp Glu Ala Lys 100 105 110 Glu Lys Asp Ile Asn Tyr Ala Ala Pro Leu Tyr Val Thr Ala Glu Phe 115 120 125 Val Asn Asn Thr Thr Gly Glu Ile Lys Ser Gln Thr Val Phe Ile Gly 130 135 140 Asp Phe Pro Met Met Thr Asp Lys Gly Thr Phe Ile Ile Asn Gly Thr 145 150 155 160 Glu Arg Val Val Val Ser Gln Leu Val Arg Ser Pro Gly Val Tyr Phe 165 170 175 Asp Gln Thr Ile Asp Lys Ser Thr Glu Arg Pro Leu His Ala Val Lys 180 185 190 Val Ile Pro Ser Arg Gly Ala Trp Leu Glu Phe Asp Val Asp Lys Arg 195 200 205 Asp Ser Val Gly Val Arg Ile Asp Arg Lys Arg Arg Gln Pro Val Thr 210 215 220 Val Leu Leu Lys Ala Leu Gly Trp Thr Thr Glu Gln Ile Thr Glu Arg 225 230 235 240 Phe Gly Phe Ser Glu Ile Met Met Ser Thr Leu Glu Ser Asp Gly Val 245 250 255 Ala Asn Thr Asp Glu Ala Leu Leu Glu Ile Tyr Arg Lys Gln Arg Pro 260 265 270 Gly Glu Gln Pro Thr Arg Asp Leu Ala Gln Ser Leu Leu Asp Asn Ser 275 280 285 Phe Phe Arg Ala Lys Arg Tyr Asp Leu Ala Arg Val Gly Arg Tyr Lys 290 295 300 Ile Asn Arg Lys Leu Gly Leu Gly Gly Asp His Asp Gly Leu Met Thr 305 310 315 320 Leu Thr Glu Glu Asp Ile Ala Thr Thr Ile Glu Tyr Leu Val Arg Leu 325 330 335 His Ala Gly Glu Arg Val Met Thr Ser Pro Asn Gly Glu Glu Ile Pro 340 345 350 Val Glu Thr Asp Asp Ile Asp His Phe Gly Asn Arg Arg Leu Arg Thr 355 360 365 Val Gly Glu Leu Ile Gln Asn Gln Val Arg Val Gly Leu Ser Arg Met 370 375 380 Glu Arg Val Val Arg Glu Arg Met Thr Thr Gln Asp Ala Glu Ser Ile 385 390 395 400 Thr Pro Thr Ser Leu Ile Asn Val Arg Pro Val Ser Ala Ala Ile Arg 405 410 415 Glu Phe Phe Gly Thr Ser Gln Leu Ser Gln Phe Met Asp Gln Asn Asn 420 425 430 Ser Leu Ser Gly Leu Thr His Lys Arg Arg Leu Ser Ala Leu Gly Pro 435 440 445 Gly Gly Leu Ser Arg Glu Arg Ala Gly Ile Glu Val Arg Asp Val His 450 455 460 Pro Ser His Tyr Gly Arg Met Cys Pro Ile Glu Thr Pro Glu Gly Pro 465 470 475 480 Asn Ile Gly Leu Ile Gly Ser Leu Ala Ser Tyr Ala Arg Val Asn Pro 485 490 495 Phe Gly Phe Ile Glu Thr Pro Tyr Arg Arg Ile Ile Asp Gly Lys Leu 500 505 510 Thr Asp Gln Ile Asp Tyr Leu Thr Ala Asp Glu Glu Asp Arg Phe Val 515 520 525 Val Ala Gln Ala Asn Thr His Tyr Asp Glu Glu Gly Asn Ile Thr Asp 530 535 540 Glu Thr Val Thr Val Arg Leu Lys Asp Gly Asp Ile Ala Met Val Gly 545 550 555 560 Arg Asn Ala Val Asp Tyr Met Asp Val Ser Pro Arg Gln Met Val Ser 565 570 575 Val Gly Thr Ala Met Ile Pro Phe Leu Glu His Asp Asp Ala Asn Arg 580 585 590 Ala Leu Met Gly Ala Asn Met Gln Lys Gln Ala Val Pro Leu Ile Arg 595 600 605 Ala Glu Ala Pro Phe Val Gly Thr Gly Met Glu Gln Arg Ala Ala Tyr 610 615 620 Asp Ala Gly Asp Leu Val Ile Thr Pro Val Ala Gly Val Val Glu Asn 625 630 635 640 Val Ser Ala Asp Phe Ile Thr Ile Met Ala Asp Asp Gly Lys Arg Glu 645 650 655 Thr Tyr Leu Leu Arg Lys Phe Gln Arg Thr Asn Gln Gly Thr Ser Tyr 660 665 670 Asn Gln Lys Pro Leu Val Asn Leu Gly Glu Arg Val Glu Ala Gly Gln 675 680 685 Val Ile Ala Asp Gly Pro Gly Thr Phe Asn Gly Glu Met Ser Leu Gly 690 695 700 Arg Asn Leu Leu Val Ala Phe Met Pro Trp Glu Gly His Asn Tyr Glu 705 710 715 720 Asp Ala Ile Ile Leu Asn Gln Asn Ile Val Glu Gln Asp Ile Leu Thr 725 730 735 Ser Ile His Ile Glu Glu His Glu Ile Asp Ala Arg Asp Thr Lys Leu 740 745 750 Gly Ala Glu Glu Ile Thr Arg Asp Ile Pro Asn Val Ser Glu Glu Val 755 760 765 Leu Lys Asp Leu Asp Asp Arg Gly Ile Val Arg Ile Gly Ala Asp Val 770 775 780 Arg Asp Gly Asp Ile Leu Val Gly Lys Val Thr Pro Lys Gly Glu Thr 785 790 795 800 Glu Leu Thr Pro Glu Glu Arg Leu Leu Arg Ala Ile Phe Gly Glu Lys 805 810 815 Ala Arg Glu Val Arg Asp Thr Ser Met Lys Val Pro His Gly Glu Thr 820 825 830 Gly Lys Val Ile Gly Val Arg His Phe Ser Arg Glu Asp Asp Asp Asp 835 840 845 Leu Ala Pro Gly Val Asn Glu Met Ile Arg Ile Tyr Val Ala Gln Lys 850 855 860 Arg Lys Ile Gln Asp Gly Asp Lys Leu Ala Gly Arg His Gly Asn Lys 865 870 875 880 Gly Val Val Gly Lys Ile Leu Pro Gln Glu Asp Met Pro Phe Leu Pro 885 890 895 Asp Gly Thr Pro Val Asp Ile Ile Leu Asn Thr His Gly Val Pro Arg 900 905 910 Arg Met Asn Ile Gly Gln Val Leu Glu Thr His Leu Gly Trp Leu Ala 915 920 925 Ser Ala Gly Trp Ser Val Asp Pro Glu Asp Pro Glu Asn Ala Glu Leu 930 935 940 Val Lys Thr Leu Pro Ala Asp Leu Leu Glu Val Pro Ala Gly Ser Leu 945 950 955 960 Thr Ala Thr Pro Val Phe Asp Gly Ala Ser Asn Glu Glu Leu Ala Gly 965 970 975 Leu Leu Ala Asn Ser Arg Pro Asn Arg Asp Gly Asp Val Met Val Asn 980 985 990 Ala Asp Gly Lys Ala Thr Leu Ile Asp Gly Arg Ser Gly Glu Pro Tyr 995 1000 1005 Pro Tyr Pro Val Ser Ile Gly Tyr Met Tyr Met Leu Lys Leu His 1010 1015 1020 His Leu Val Asp Glu Lys Ile His Ala Arg Ser Thr Gly Pro Tyr 1025 1030 1035 Ser Met Ile Thr Gln Gln Pro Leu Gly Gly Lys Ala Gln Phe Gly 1040 1045 1050 Gly Gln Arg Phe Gly Glu Met Glu Val Trp Ala Met Gln Ala Tyr 1055 1060 1065 Gly Ala Ala Tyr Thr Leu Gln Glu Leu Leu Thr Ile Lys Ser Asp 1070 1075 1080 Asp Val Val Gly Arg Val Lys Val Tyr Glu Ala Ile Val Lys Gly 1085 1090 1095 Glu Asn Ile Pro Asp Pro Gly Ile Pro Glu Ser Phe Lys Val Leu 1100 1105 1110 Leu Lys Glu Leu Gln Ser Leu Cys Leu Asn Val Glu Val Leu Ser 1115 1120 1125 Ala Asp Gly Thr Pro Met Glu Leu Ala Gly Asp Asp Asp Asp Phe 1130 1135 1140 Asp Gln Ala Gly Ala Ser Leu Gly Ile Asn Leu Ser Arg Asp Glu 1145 1150 1155 Arg Ser Asp Ala Asp Thr Ala 1160 1165 3 5099 DNA Corynebacterium glutamicum CDS (702)..(4196) 3 acaatgtgac tcgtgatttt tgggtggatc agcgtaccgg tttggttgtc gatctagctg 60 aaaatattga tgatttttac ggcgaccgca gcggccagaa gtacgaacag aaattgcttt 120 tcgacgcctc cctcgacgat gcagctgtct ctaagctggt tgcacaggcc gaaagcatcc 180 ctgatggaga tgtgagcaaa atcgcaaata ccgtaggtat tgtgatcggt gcggtattgg 240 ctctcgtggg cctggccggg tgttttgggg cgtttgggaa gaaacgtcga gaagcttaac 300 ctgctgttca aatagatttt ccctgtttcg aattgcggaa accccgggtt tgtttgctag 360 ggtgcctcgt agaaggggtc aagaagattt ctgggaaacg cgcccgtgcg gttggttgct 420 aatagcacgc ggagcaccag atgaaaaatc tcccctttac tttcgcgcgc gattggtata 480 ctctgagtcg ttgcgttgga attcgtgact ctttttcgtt cctgtagcgc caagaccttg 540 atcaaggtgg tttaaaaaaa ccgatttgac aaggtcattc agtgctatct ggagtcgttc 600 agggggatcg ggttcctcag cagaccaatt gctcaaaaat accagcggtg ttgatctgca 660 cttaatggcc ttgaccagcc aggtgcaatt acccgcgtga g gtg ctg gaa gga ctc 716 Val Leu Glu Gly Leu 1 5 atc ttg gca gtc tcc cgc cag acc aag tca gtc gtc gat att ccc ggt 764 Ile Leu Ala Val Ser Arg Gln Thr Lys Ser Val Val Asp Ile Pro Gly 10 15 20 gca ccg cag cgt tat tct ttc gcg aag gtg tcc gca ccc att gag gtg 812 Ala Pro Gln Arg Tyr Ser Phe Ala Lys Val Ser Ala Pro Ile Glu Val 25 30 35 ccc ggg cta cta gat ctt caa ctg gat tct tac tcc tgg ctg att ggt 860 Pro Gly Leu Leu Asp Leu Gln Leu Asp Ser Tyr Ser Trp Leu Ile Gly 40 45 50 acg cct gag tgg cgt gct cgt cag aag gaa gaa ttc ggc gag gga gcc 908 Thr Pro Glu Trp Arg Ala Arg Gln Lys Glu Glu Phe Gly Glu Gly Ala 55 60 65 cgc gta acc agc ggc ctt gag aac att ctc gag gag ctc tcc cca atc 956 Arg Val Thr Ser Gly Leu Glu Asn Ile Leu Glu Glu Leu Ser Pro Ile 70 75 80 85 cag gat tac tct gga aac atg tcc ctg agc ctt tcg gag cca cgc ttc 1004 Gln Asp Tyr Ser Gly Asn Met Ser Leu Ser Leu Ser Glu Pro Arg Phe 90 95 100 gaa gac gtc aag aac acc att gac gag gcg aaa gaa aag gac atc aac 1052 Glu Asp Val Lys Asn Thr Ile Asp Glu Ala Lys Glu Lys Asp Ile Asn 105 110 115 tac gcg gcg cca ctg tat gtg acc gcg gag ttc gtc aac aac acc acc 1100 Tyr Ala Ala Pro Leu Tyr Val Thr Ala Glu Phe Val Asn Asn Thr Thr 120 125 130 ggt gaa atc aag tct cag act gtc ttc atc ggc gat ttc cca atg atg 1148 Gly Glu Ile Lys Ser Gln Thr Val Phe Ile Gly Asp Phe Pro Met Met 135 140 145 acg gac aag gga acg ttc atc atc aac gga acc gaa cgc gtt gtg gtc 1196 Thr Asp Lys Gly Thr Phe Ile Ile Asn Gly Thr Glu Arg Val Val Val 150 155 160 165 agc cag ctc gtc cgc tcc ccg ggc gtg tac ttt gac cag acc atc gat 1244 Ser Gln Leu Val Arg Ser Pro Gly Val Tyr Phe Asp Gln Thr Ile Asp 170 175 180 aag tca act gag cgt cca ctg cac gcc gtg aag gtt att cct ttc cgt 1292 Lys Ser Thr Glu Arg Pro Leu His Ala Val Lys Val Ile Pro Phe Arg 185 190 195 ggt gct tgg ctt gag ttt gac gtc gat aag cgc gat tcg gtt ggt gtt 1340 Gly Ala Trp Leu Glu Phe Asp Val Asp Lys Arg Asp Ser Val Gly Val 200 205 210 cgt att gac cgc aag cgt cgc cag cca gtc acc gta ctg ctg aag gct 1388 Arg Ile Asp Arg Lys Arg Arg Gln Pro Val Thr Val Leu Leu Lys Ala 215 220 225 ctt ggc tgg acc act gag cag atc acc gag cgt ttc ggt ttc tct gaa 1436 Leu Gly Trp Thr Thr Glu Gln Ile Thr Glu Arg Phe Gly Phe Ser Glu 230 235 240 245 atc atg atg tcc acc ctc gag tcc gat ggt gta gca aac acc gat gag 1484 Ile Met Met Ser Thr Leu Glu Ser Asp Gly Val Ala Asn Thr Asp Glu 250 255 260 gca ttg ctg gag atc tac cgc aag cag cgt cca ggc gag cag cct acc 1532 Ala Leu Leu Glu Ile Tyr Arg Lys Gln Arg Pro Gly Glu Gln Pro Thr 265 270 275 cgc gac ctt gcg cag tcc ctc ctg gac aac agc ttc ttc cgt gca aag 1580 Arg Asp Leu Ala Gln Ser Leu Leu Asp Asn Ser Phe Phe Arg Ala Lys 280 285 290 cgc tac gac ctg gct cgc gtt ggt cgt tac aag atc aac cgc aag ctc 1628 Arg Tyr Asp Leu Ala Arg Val Gly Arg Tyr Lys Ile Asn Arg Lys Leu 295 300 305 ggc ctt ggt ggc gac cac gat ggt ttg atg act ctt act gaa gag gac 1676 Gly Leu Gly Gly Asp His Asp Gly Leu Met Thr Leu Thr Glu Glu Asp 310 315 320 325 atc gca acc acc atc gag tac ctg gtg cgt ctg cac gca ggt gag cgc 1724 Ile Ala Thr Thr Ile Glu Tyr Leu Val Arg Leu His Ala Gly Glu Arg 330 335 340 gtc atg act tct cca aat ggt gaa gag atc cca gtc gag acc gat gac 1772 Val Met Thr Ser Pro Asn Gly Glu Glu Ile Pro Val Glu Thr Asp Asp 345 350 355 atc gac cac ttt ggt aac cgt cgt ctg cgt acc gtt ggc gaa ctg atc 1820 Ile Asp His Phe Gly Asn Arg Arg Leu Arg Thr Val Gly Glu Leu Ile 360 365 370 cag aac cag gtc cgt gtc ggc ctg tcc cgc atg gag cgc gtt gtt cgt 1868 Gln Asn Gln Val Arg Val Gly Leu Ser Arg Met Glu Arg Val Val Arg 375 380 385 gag cgt atg acc acc cag gat gcg gag tcc att act cct act tcc ttg 1916 Glu Arg Met Thr Thr Gln Asp Ala Glu Ser Ile Thr Pro Thr Ser Leu 390 395 400 405 atc aac gtt cgt cct gtc tct gca gct atc cgt gag ttc ttc gga act 1964 Ile Asn Val Arg Pro Val Ser Ala Ala Ile Arg Glu Phe Phe Gly Thr 410 415 420 tcc cag ctg tct cag ttc atg gtc cag aac aac tcc ctg tct ggt ttg 2012 Ser Gln Leu Ser Gln Phe Met Val Gln Asn Asn Ser Leu Ser Gly Leu 425 430 435 act cac aag cgt cgt ctg tcg gct ctg ggc ccg ggt ggt ctg tcc cgt 2060 Thr His Lys Arg Arg Leu Ser Ala Leu Gly Pro Gly Gly Leu Ser Arg 440 445 450 gag cgc gcc ggc atc gag gtt cga gac gtt cac cca tct cac tac ggc 2108 Glu Arg Ala Gly Ile Glu Val Arg Asp Val His Pro Ser His Tyr Gly 455 460 465 cgt atg tgc cca att gag act ccg gaa ggt cca aac att ggc ctg atc 2156 Arg Met Cys Pro Ile Glu Thr Pro Glu Gly Pro Asn Ile Gly Leu Ile 470 475 480 485 ggt tcc ttg gct tcc tat gct cga gtg aac cca ttc ggt ttc att gag 2204 Gly Ser Leu Ala Ser Tyr Ala Arg Val Asn Pro Phe Gly Phe Ile Glu 490 495 500 acc cca tac cgt cgc atc atc gac ggc aag ctg acc gac cag att gac 2252 Thr Pro Tyr Arg Arg Ile Ile Asp Gly Lys Leu Thr Asp Gln Ile Asp 505 510 515 tac ctt acc gct gat gag gaa gac cgc ttc gtt gtt gcg cag gca aac 2300 Tyr Leu Thr Ala Asp Glu Glu Asp Arg Phe Val Val Ala Gln Ala Asn 520 525 530 acg cac tac gac gaa gag ggc aac atc acc gat gag acc gtc act gtt 2348 Thr His Tyr Asp Glu Glu Gly Asn Ile Thr Asp Glu Thr Val Thr Val 535 540 545 cgt ctg aag gac ggc gac atc gcc atg gtt ggc cgc aac gcg gtt gat 2396 Arg Leu Lys Asp Gly Asp Ile Ala Met Val Gly Arg Asn Ala Val Asp 550 555 560 565 tac atg gac gtt tcc cct cgt cag atg gtt tct gtt ggt acc gcg atg 2444 Tyr Met Asp Val Ser Pro Arg Gln Met Val Ser Val Gly Thr Ala Met 570 575 580 att cca ttc ctg gag cac gac gat gct aac cgt gca ctg atg ggc gcg 2492 Ile Pro Phe Leu Glu His Asp Asp Ala Asn Arg Ala Leu Met Gly Ala 585 590 595 aac atg cag aag cag gct gtg cca ctg att cgt gcc gag gct cct ttc 2540 Asn Met Gln Lys Gln Ala Val Pro Leu Ile Arg Ala Glu Ala Pro Phe 600 605 610 gtg ggc acc ggt atg gag cag cgc gca gca tac gac gcc ggc gac ctg 2588 Val Gly Thr Gly Met Glu Gln Arg Ala Ala Tyr Asp Ala Gly Asp Leu 615 620 625 gtt att acc cca gtc gca ggt gtg gtg gaa aac gtt tca gct gac ttc 2636 Val Ile Thr Pro Val Ala Gly Val Val Glu Asn Val Ser Ala Asp Phe 630 635 640 645 atc acc atc atg gct gat gac ggc aag cgc gaa acc tac ctg ctg cgt 2684 Ile Thr Ile Met Ala Asp Asp Gly Lys Arg Glu Thr Tyr Leu Leu Arg 650 655 660 aag ttc cag cgc acc aac cag ggc acc agc tac aac cag aag cct ttg 2732 Lys Phe Gln Arg Thr Asn Gln Gly Thr Ser Tyr Asn Gln Lys Pro Leu 665 670 675 gtt aac ttg ggc gag cgc gtt gaa gct ggc cag gtt att gct gat ggt 2780 Val Asn Leu Gly Glu Arg Val Glu Ala Gly Gln Val Ile Ala Asp Gly 680 685 690 cca ggt acc ttc aat ggt gaa atg tcc ctt ggc cgt aac ctt ctg gtt 2828 Pro Gly Thr Phe Asn Gly Glu Met Ser Leu Gly Arg Asn Leu Leu Val 695 700 705 gcg ttc atg cct tgg gaa ggc cac aac tac gag gat gcg atc atc ctc 2876 Ala Phe Met Pro Trp Glu Gly His Asn Tyr Glu Asp Ala Ile Ile Leu 710 715 720 725 aac cag aac atc gtt gag cag gac atc ttg acc tcg atc cac atc gag 2924 Asn Gln Asn Ile Val Glu Gln Asp Ile Leu Thr Ser Ile His Ile Glu 730 735 740 gag cac gag atc gat gcc cgc gac act aag ctt ggc gcc gaa gaa atc 2972 Glu His Glu Ile Asp Ala Arg Asp Thr Lys Leu Gly Ala Glu Glu Ile 745 750 755 acc cgc gac atc cct aat gtg tct gaa gaa gtc ctc aag gac ctc gac 3020 Thr Arg Asp Ile Pro Asn Val Ser Glu Glu Val Leu Lys Asp Leu Asp 760 765 770 gac cgc ggt att gtc cgc atc ggt gct gat gtt cgt gac ggc gac atc 3068 Asp Arg Gly Ile Val Arg Ile Gly Ala Asp Val Arg Asp Gly Asp Ile 775 780 785 ctg gtc ggt aag gtc acc cct aag ggc gag acc gag ctc acc ccg gaa 3116 Leu Val Gly Lys Val Thr Pro Lys Gly Glu Thr Glu Leu Thr Pro Glu 790 795 800 805 gag cgc ttg ctg cgc gca atc ttc ggt gag aag gcc cgc gaa gtt cgc 3164 Glu Arg Leu Leu Arg Ala Ile Phe Gly Glu Lys Ala Arg Glu Val Arg 810 815 820 gat acc tcc atg aag gtg cct cac ggt gag acc ggc aag gtc atc ggc 3212 Asp Thr Ser Met Lys Val Pro His Gly Glu Thr Gly Lys Val Ile Gly 825 830 835 gtg cgt cac ttc tcc cgc gag gac gac gac gat ctg gct cct ggc gtc 3260 Val Arg His Phe Ser Arg Glu Asp Asp Asp Asp Leu Ala Pro Gly Val 840 845 850 aac gag atg atc cgt atc tac gtt gct cag aag cgt aag atc cag gac 3308 Asn Glu Met Ile Arg Ile Tyr Val Ala Gln Lys Arg Lys Ile Gln Asp 855 860 865 ggc gat aag ctc gct ggc cgc cac ggt aac aag ggt gtt gtc ggt aaa 3356 Gly Asp Lys Leu Ala Gly Arg His Gly Asn Lys Gly Val Val Gly Lys 870 875 880 885 att ttg cct cag gaa gat atg cca ttc ctt cca gac ggc act cct gtt 3404 Ile Leu Pro Gln Glu Asp Met Pro Phe Leu Pro Asp Gly Thr Pro Val 890 895 900 gac atc atc ttg aac acc cac ggt gtt cca cgt cgt atg aac att ggt 3452 Asp Ile Ile Leu Asn Thr His Gly Val Pro Arg Arg Met Asn Ile Gly 905 910 915 cag gtt ctt gag acc cac ctt ggc tgg ctg gca tct gct ggt tgg tcc 3500 Gln Val Leu Glu Thr His Leu Gly Trp Leu Ala Ser Ala Gly Trp Ser 920 925 930 gtg gat cct gaa gat cct gag aac gct gag ctc gtc aag act ctg cct 3548 Val Asp Pro Glu Asp Pro Glu Asn Ala Glu Leu Val Lys Thr Leu Pro 935 940 945 gca gac ctc ctc gag gtt cct gct ggt tcc ttg act gca act cct gtg 3596 Ala Asp Leu Leu Glu Val Pro Ala Gly Ser Leu Thr Ala Thr Pro Val 950 955 960 965 ttc gac ggt gcg tca aac gaa gag ctc gca ggc ctg ctc gct aat tca 3644 Phe Asp Gly Ala Ser Asn Glu Glu Leu Ala Gly Leu Leu Ala Asn Ser 970 975 980 cgt cca aac cgc gac ggc gac gtc atg gtt aac gcg gat ggt aaa gca 3692 Arg Pro Asn Arg Asp Gly Asp Val Met Val Asn Ala Asp Gly Lys Ala 985 990 995 acg ctt atc gac ggt cgc tcc ggt gag cct tac ccg tac ccg gtt 3737 Thr Leu Ile Asp Gly Arg Ser Gly Glu Pro Tyr Pro Tyr Pro Val 1000 1005 1010 tcc atc ggc tac atg tac atg ctg aag ctg cac cac ctc gtt gac 3782 Ser Ile Gly Tyr Met Tyr Met Leu Lys Leu His His Leu Val Asp 1015 1020 1025 gag aag atc cac gca cgt tcc act ggt cct tac tcc atg att acc 3827 Glu Lys Ile His Ala Arg Ser Thr Gly Pro Tyr Ser Met Ile Thr 1030 1035 1040 cag cag cca ctg ggt ggt aaa gca cag ttc ggt gga cag cgt ttc 3872 Gln Gln Pro Leu Gly Gly Lys Ala Gln Phe Gly Gly Gln Arg Phe 1045 1050 1055 ggc gaa atg gag gtg tgg gca atg cag gca tac ggc gct gcc tac 3917 Gly Glu Met Glu Val Trp Ala Met Gln Ala Tyr Gly Ala Ala Tyr 1060 1065 1070 aca ctt cag gag ctg ctg acc atc aag tct gat gac gtg gtt ggc 3962 Thr Leu Gln Glu Leu Leu Thr Ile Lys Ser Asp Asp Val Val Gly 1075 1080 1085 cgt gtc aag gtc tac gaa gca att gtg aag ggc gag aac atc ccg 4007 Arg Val Lys Val Tyr Glu Ala Ile Val Lys Gly Glu Asn Ile Pro 1090 1095 1100 gat cca ggt att cct gag tcc ttc aag gtt ctc ctc aag gag ctc 4052 Asp Pro Gly Ile Pro Glu Ser Phe Lys Val Leu Leu Lys Glu Leu 1105 1110 1115 cag tcc ttg tgc ctg aac gtg gag gtt ctc tcc gca gac ggc act 4097 Gln Ser Leu Cys Leu Asn Val Glu Val Leu Ser Ala Asp Gly Thr 1120 1125 1130 cca atg gag ctc gcg ggt gac gac gac gac ttc gat cag gca ggc 4142 Pro Met Glu Leu Ala Gly Asp Asp Asp Asp Phe Asp Gln Ala Gly 1135 1140 1145 gcc tca ctt ggc atc aac ctg tcc cgt gac gag cgt tcc gac gcc 4187 Ala Ser Leu Gly Ile Asn Leu Ser Arg Asp Glu Arg Ser Asp Ala 1150 1155 1160 gac acc gca tagcagatca gaaaacaacc gctagaaatc aagccataca 4236 Asp Thr Ala 1165 tcccccggac attgaagaga tgttctgggg ggaaagggag ttttacgtgc tcgacgtaaa 4296 cgtcttcgat gagctccgca tcggcctggc caccgccgac gacatccgcc gttggtccaa 4356 gggtgaggtc aagaagccgg agaccatcaa ctaccgaacc ctcaagcctg agaaggacgg 4416 tctgttctgc gagcgtatct tcggtccaac tcgcgactgg gagtgcgcct gcggtaagta 4476 caagcgtgtc cgctacaagg gcatcatctg tgaacgctgt ggcgttgagg tcaccaagtc 4536 caaggtgcgc cgtgagcgca tgggacacat tgagctcgct gcaccagtaa cccacatttg 4596 gtacttcaag ggcgttccat cacgcctcgg ctaccttttg gaccttgctc caaaggacct 4656 ggacctcatc atctacttcg gtgcgaacat catcaccagc gtggacgaag aggctcgcca 4716 cagcgaccag accactcttg aggcagaaat gcttctggag aagaaggacg ttgaggcaga 4776 cgcagagtct gacattgctg agcgtgctga aaagctcgaa gaggatcttg ctgaacttga 4836 ggcagctggc gctaaggccg acgctcgccg caaggttcag gctgctgccg ataaggaaat 4896 gcagcacatc cgtgagcgtg cacagcgcga aatcgatcgt ctcgatgagg tctggcagac 4956 cttcatcaag cttgctccaa agcagatgat ccgcgatgag aagctctacg atgaactgat 5016 cgaccgctac gaggattact tcaccggtgg tatgggtgca gagtccattg aggctttgat 5076 ccagaacttc gaccttgatg ctg 5099 4 1165 PRT Corynebacterium glutamicum 4 Val Leu Glu Gly Leu Ile Leu Ala Val Ser Arg Gln Thr Lys Ser Val 1 5 10 15 Val Asp Ile Pro Gly Ala Pro Gln Arg Tyr Ser Phe Ala Lys Val Ser 20 25 30 Ala Pro Ile Glu Val Pro Gly Leu Leu Asp Leu Gln Leu Asp Ser Tyr 35 40 45 Ser Trp Leu Ile Gly Thr Pro Glu Trp Arg Ala Arg Gln Lys Glu Glu 50 55 60 Phe Gly Glu Gly Ala Arg Val Thr Ser Gly Leu Glu Asn Ile Leu Glu 65 70 75 80 Glu Leu Ser Pro Ile Gln Asp Tyr Ser Gly Asn Met Ser Leu Ser Leu 85 90 95 Ser Glu Pro Arg Phe Glu Asp Val Lys Asn Thr Ile Asp Glu Ala Lys 100 105 110 Glu Lys Asp Ile Asn Tyr Ala Ala Pro Leu Tyr Val Thr Ala Glu Phe 115 120 125 Val Asn Asn Thr Thr Gly Glu Ile Lys Ser Gln Thr Val Phe Ile Gly 130 135 140 Asp Phe Pro Met Met Thr Asp Lys Gly Thr Phe Ile Ile Asn Gly Thr 145 150 155 160 Glu Arg Val Val Val Ser Gln Leu Val Arg Ser Pro Gly Val Tyr Phe 165 170 175 Asp Gln Thr Ile Asp Lys Ser Thr Glu Arg Pro Leu His Ala Val Lys 180 185 190 Val Ile Pro Phe Arg Gly Ala Trp Leu Glu Phe Asp Val Asp Lys Arg 195 200 205 Asp Ser Val Gly Val Arg Ile Asp Arg Lys Arg Arg Gln Pro Val Thr 210 215 220 Val Leu Leu Lys Ala Leu Gly Trp Thr Thr Glu Gln Ile Thr Glu Arg 225 230 235 240 Phe Gly Phe Ser Glu Ile Met Met Ser Thr Leu Glu Ser Asp Gly Val 245 250 255 Ala Asn Thr Asp Glu Ala Leu Leu Glu Ile Tyr Arg Lys Gln Arg Pro 260 265 270 Gly Glu Gln Pro Thr Arg Asp Leu Ala Gln Ser Leu Leu Asp Asn Ser 275 280 285 Phe Phe Arg Ala Lys Arg Tyr Asp Leu Ala Arg Val Gly Arg Tyr Lys 290 295 300 Ile Asn Arg Lys Leu Gly Leu Gly Gly Asp His Asp Gly Leu Met Thr 305 310 315 320 Leu Thr Glu Glu Asp Ile Ala Thr Thr Ile Glu Tyr Leu Val Arg Leu 325 330 335 His Ala Gly Glu Arg Val Met Thr Ser Pro Asn Gly Glu Glu Ile Pro 340 345 350 Val Glu Thr Asp Asp Ile Asp His Phe Gly Asn Arg Arg Leu Arg Thr 355 360 365 Val Gly Glu Leu Ile Gln Asn Gln Val Arg Val Gly Leu Ser Arg Met 370 375 380 Glu Arg Val Val Arg Glu Arg Met Thr Thr Gln Asp Ala Glu Ser Ile 385 390 395 400 Thr Pro Thr Ser Leu Ile Asn Val Arg Pro Val Ser Ala Ala Ile Arg 405 410 415 Glu Phe Phe Gly Thr Ser Gln Leu Ser Gln Phe Met Val Gln Asn Asn 420 425 430 Ser Leu Ser Gly Leu Thr His Lys Arg Arg Leu Ser Ala Leu Gly Pro 435 440 445 Gly Gly Leu Ser Arg Glu Arg Ala Gly Ile Glu Val Arg Asp Val His 450 455 460 Pro Ser His Tyr Gly Arg Met Cys Pro Ile Glu Thr Pro Glu Gly Pro 465 470 475 480 Asn Ile Gly Leu Ile Gly Ser Leu Ala Ser Tyr Ala Arg Val Asn Pro 485 490 495 Phe Gly Phe Ile Glu Thr Pro Tyr Arg Arg Ile Ile Asp Gly Lys Leu 500 505 510 Thr Asp Gln Ile Asp Tyr Leu Thr Ala Asp Glu Glu Asp Arg Phe Val 515 520 525 Val Ala Gln Ala Asn Thr His Tyr Asp Glu Glu Gly Asn Ile Thr Asp 530 535 540 Glu Thr Val Thr Val Arg Leu Lys Asp Gly Asp Ile Ala Met Val Gly 545 550 555 560 Arg Asn Ala Val Asp Tyr Met Asp Val Ser Pro Arg Gln Met Val Ser 565 570 575 Val Gly Thr Ala Met Ile Pro Phe Leu Glu His Asp Asp Ala Asn Arg 580 585 590 Ala Leu Met Gly Ala Asn Met Gln Lys Gln Ala Val Pro Leu Ile Arg 595 600 605 Ala Glu Ala Pro Phe Val Gly Thr Gly Met Glu Gln Arg Ala Ala Tyr 610 615 620 Asp Ala Gly Asp Leu Val Ile Thr Pro Val Ala Gly Val Val Glu Asn 625 630 635 640 Val Ser Ala Asp Phe Ile Thr Ile Met Ala Asp Asp Gly Lys Arg Glu 645 650 655 Thr Tyr Leu Leu Arg Lys Phe Gln Arg Thr Asn Gln Gly Thr Ser Tyr 660 665 670 Asn Gln Lys Pro Leu Val Asn Leu Gly Glu Arg Val Glu Ala Gly Gln 675 680 685 Val Ile Ala Asp Gly Pro Gly Thr Phe Asn Gly Glu Met Ser Leu Gly 690 695 700 Arg Asn Leu Leu Val Ala Phe Met Pro Trp Glu Gly His Asn Tyr Glu 705 710 715 720 Asp Ala Ile Ile Leu Asn Gln Asn Ile Val Glu Gln Asp Ile Leu Thr 725 730 735 Ser Ile His Ile Glu Glu His Glu Ile Asp Ala Arg Asp Thr Lys Leu 740 745 750 Gly Ala Glu Glu Ile Thr Arg Asp Ile Pro Asn Val Ser Glu Glu Val 755 760 765 Leu Lys Asp Leu Asp Asp Arg Gly Ile Val Arg Ile Gly Ala Asp Val 770 775 780 Arg Asp Gly Asp Ile Leu Val Gly Lys Val Thr Pro Lys Gly Glu Thr 785 790 795 800 Glu Leu Thr Pro Glu Glu Arg Leu Leu Arg Ala Ile Phe Gly Glu Lys 805 810 815 Ala Arg Glu Val Arg Asp Thr Ser Met Lys Val Pro His Gly Glu Thr 820 825 830 Gly Lys Val Ile Gly Val Arg His Phe Ser Arg Glu Asp Asp Asp Asp 835 840 845 Leu Ala Pro Gly Val Asn Glu Met Ile Arg Ile Tyr Val Ala Gln Lys 850 855 860 Arg Lys Ile Gln Asp Gly Asp Lys Leu Ala Gly Arg His Gly Asn Lys 865 870 875 880 Gly Val Val Gly Lys Ile Leu Pro Gln Glu Asp Met Pro Phe Leu Pro 885 890 895 Asp Gly Thr Pro Val Asp Ile Ile Leu Asn Thr His Gly Val Pro Arg 900 905 910 Arg Met Asn Ile Gly Gln Val Leu Glu Thr His Leu Gly Trp Leu Ala 915 920 925 Ser Ala Gly Trp Ser Val Asp Pro Glu Asp Pro Glu Asn Ala Glu Leu 930 935 940 Val Lys Thr Leu Pro Ala Asp Leu Leu Glu Val Pro Ala Gly Ser Leu 945 950 955 960 Thr Ala Thr Pro Val Phe Asp Gly Ala Ser Asn Glu Glu Leu Ala Gly 965 970 975 Leu Leu Ala Asn Ser Arg Pro Asn Arg Asp Gly Asp Val Met Val Asn 980 985 990 Ala Asp Gly Lys Ala Thr Leu Ile Asp Gly Arg Ser Gly Glu Pro Tyr 995 1000 1005 Pro Tyr Pro Val Ser Ile Gly Tyr Met Tyr Met Leu Lys Leu His 1010 1015 1020 His Leu Val Asp Glu Lys Ile His Ala Arg Ser Thr Gly Pro Tyr 1025 1030 1035 Ser Met Ile Thr Gln Gln Pro Leu Gly Gly Lys Ala Gln Phe Gly 1040 1045 1050 Gly Gln Arg Phe Gly Glu Met Glu Val Trp Ala Met Gln Ala Tyr 1055 1060 1065 Gly Ala Ala Tyr Thr Leu Gln Glu Leu Leu Thr Ile Lys Ser Asp 1070 1075 1080 Asp Val Val Gly Arg Val Lys Val Tyr Glu Ala Ile Val Lys Gly 1085 1090 1095 Glu Asn Ile Pro Asp Pro Gly Ile Pro Glu Ser Phe Lys Val Leu 1100 1105 1110 Leu Lys Glu Leu Gln Ser Leu Cys Leu Asn Val Glu Val Leu Ser 1115 1120 1125 Ala Asp Gly Thr Pro Met Glu Leu Ala Gly Asp Asp Asp Asp Phe 1130 1135 1140 Asp Gln Ala Gly Ala Ser Leu Gly Ile Asn Leu Ser Arg Asp Glu 1145 1150 1155 Arg Ser Asp Ala Asp Thr Ala 1160 1165 5 5099 DNA Corynebacterium glutamicum CDS (702)..(4196) 5 acaatgtgac tcgtgatttt tgggtggatc agcgtaccgg tttggttgtc gatctagctg 60 aaaatattga tgatttttac ggcgaccgca gcggccagaa gtacgaacag aaattgcttt 120 tcgacgcctc cctcgacgat gcagctgtct ctaagctggt tgcacaggcc gaaagcatcc 180 ctgatggaga tgtgagcaaa atcgcaaata ccgtaggtat tgtgatcggt gcggtattgg 240 ctctcgtggg cctggccggg tgttttgggg cgtttgggaa gaaacgtcga gaagcttaac 300 ctgctgttca aatagatttt ccctgtttcg aattgcggaa accccgggtt tgtttgctag 360 ggtgcctcgt agaaggggtc aagaagattt ctgggaaacg cgcccgtgcg gttggttgct 420 aatagcacgc ggagcaccag atgaaaaatc tcccctttac tttcgcgcgc gattggtata 480 ctctgagtcg ttgcgttgga attcgtgact ctttttcgtt cctgtagcgc caagaccttg 540 atcaaggtgg tttaaaaaaa ccgatttgac aaggtcattc agtgctatct ggagtcgttc 600 agggggatcg ggttcctcag cagaccaatt gctcaaaaat accagcggtg ttgatctgca 660 cttaatggcc ttgaccagcc aggtgcaatt acccgcgtga g gtg ctg gaa gga ccc 716 Val Leu Glu Gly Pro 1 5 atc ttg gca gtc tcc cgc cag acc aag tca gtc gtc gat att ccc ggt 764 Ile Leu Ala Val Ser Arg Gln Thr Lys Ser Val Val Asp Ile Pro Gly 10 15 20 gca ccg cag cgt tat tct ttc gcg aag gtg tcc gca ccc att gag gtg 812 Ala Pro Gln Arg Tyr Ser Phe Ala Lys Val Ser Ala Pro Ile Glu Val 25 30 35 ccc ggg cta cta gat ctt caa ctg gat tct tac tcc tgg ctg att ggt 860 Pro Gly Leu Leu Asp Leu Gln Leu Asp Ser Tyr Ser Trp Leu Ile Gly 40 45 50 acg cct gag tgg cgt gct cgt cag aag gaa gaa ttc ggc gag gga gcc 908 Thr Pro Glu Trp Arg Ala Arg Gln Lys Glu Glu Phe Gly Glu Gly Ala 55 60 65 cgc gta acc agc ggc ctt gag aac att ctc gag gag ctc tcc cca atc 956 Arg Val Thr Ser Gly Leu Glu Asn Ile Leu Glu Glu Leu Ser Pro Ile 70 75 80 85 cag gat tac tct gga aac atg tcc ctg agc ctt tcg gag cca cgc ttc 1004 Gln Asp Tyr Ser Gly Asn Met Ser Leu Ser Leu Ser Glu Pro Arg Phe 90 95 100 gaa gac gtc aag aac acc att gac gag gcg aaa gaa aag gac atc aac 1052 Glu Asp Val Lys Asn Thr Ile Asp Glu Ala Lys Glu Lys Asp Ile Asn 105 110 115 tac gcg gcg cca ctg tat gtg acc gcg gag ttc gtc aac aac acc acc 1100 Tyr Ala Ala Pro Leu Tyr Val Thr Ala Glu Phe Val Asn Asn Thr Thr 120 125 130 ggt gaa atc aag tct cag act gtc ttc atc ggc gat ttc cca atg atg 1148 Gly Glu Ile Lys Ser Gln Thr Val Phe Ile Gly Asp Phe Pro Met Met 135 140 145 acg gac aag gga acg ttc atc atc aac gga acc gaa cgc gtt gtg gtc 1196 Thr Asp Lys Gly Thr Phe Ile Ile Asn Gly Thr Glu Arg Val Val Val 150 155 160 165 agc cag ctc gtc cgc tcc ccg ggc gtg tac ttt gac cag acc atc gat 1244 Ser Gln Leu Val Arg Ser Pro Gly Val Tyr Phe Asp Gln Thr Ile Asp 170 175 180 aag tca act gag cgt cca ctg cac gcc gtg aag gtt att cct tcc cgt 1292 Lys Ser Thr Glu Arg Pro Leu His Ala Val Lys Val Ile Pro Ser Arg 185 190 195 ggt gct tgg ctt gag ttt gac gtc gat aag cgc gat tcg gtt ggt gtt 1340 Gly Ala Trp Leu Glu Phe Asp Val Asp Lys Arg Asp Ser Val Gly Val 200 205 210 cgt att gac cgc aag cgt cgc cag cca gtc acc gta ctg ctg aag gct 1388 Arg Ile Asp Arg Lys Arg Arg Gln Pro Val Thr Val Leu Leu Lys Ala 215 220 225 ctt ggc tgg acc act gag cag atc acc gag cgt ttc ggt ttc tct gaa 1436 Leu Gly Trp Thr Thr Glu Gln Ile Thr Glu Arg Phe Gly Phe Ser Glu 230 235 240 245 atc atg atg tcc acc ctc gag tcc gat ggt gta gca aac acc gat gag 1484 Ile Met Met Ser Thr Leu Glu Ser Asp Gly Val Ala Asn Thr Asp Glu 250 255 260 gca ttg ctg gag atc tac cgc aag cag cgt cca ggc gag cag cct acc 1532 Ala Leu Leu Glu Ile Tyr Arg Lys Gln Arg Pro Gly Glu Gln Pro Thr 265 270 275 cgc gac ctt gcg cag tcc ctc ctg gac aac agc ttc ttc cgt gca aag 1580 Arg Asp Leu Ala Gln Ser Leu Leu Asp Asn Ser Phe Phe Arg Ala Lys 280 285 290 cgc tac gac ctg gct cgc gtt ggt cgt tac aag atc aac cgc aag ctc 1628 Arg Tyr Asp Leu Ala Arg Val Gly Arg Tyr Lys Ile Asn Arg Lys Leu 295 300 305 ggc ctt ggt ggc gac cac gat ggt ttg atg act ctt act gaa gag gac 1676 Gly Leu Gly Gly Asp His Asp Gly Leu Met Thr Leu Thr Glu Glu Asp 310 315 320 325 atc gca acc acc atc gag tac ctg gtg cgt ctg cac gca ggt gag cgc 1724 Ile Ala Thr Thr Ile Glu Tyr Leu Val Arg Leu His Ala Gly Glu Arg 330 335 340 gtc atg act tct cca aat ggt gaa gag atc cca gtc gag acc gat gac 1772 Val Met Thr Ser Pro Asn Gly Glu Glu Ile Pro Val Glu Thr Asp Asp 345 350 355 atc gac cac ttt ggt aac cgt cgt ctg cgt acc gtt ggc gaa ctg atc 1820 Ile Asp His Phe Gly Asn Arg Arg Leu Arg Thr Val Gly Glu Leu Ile 360 365 370 cag aac cag gtc cgt gtc ggc ctg tcc cgc atg gag cgc gtt gtt cgt 1868 Gln Asn Gln Val Arg Val Gly Leu Ser Arg Met Glu Arg Val Val Arg 375 380 385 gag cgt atg acc acc cag gat gcg gag tcc att act cct act tcc ttg 1916 Glu Arg Met Thr Thr Gln Asp Ala Glu Ser Ile Thr Pro Thr Ser Leu 390 395 400 405 atc aac gtt cgt cct gtc tct gca gct atc cgt gag ttc ttc gga act 1964 Ile Asn Val Arg Pro Val Ser Ala Ala Ile Arg Glu Phe Phe Gly Thr 410 415 420 tcc cag ctg tct cag ttc atg gac cag aac aac tcc ctg tct ggt ttg 2012 Ser Gln Leu Ser Gln Phe Met Asp Gln Asn Asn Ser Leu Ser Gly Leu 425 430 435 act tac aag cgt cgt ctg tcg gct ctg ggc ccg ggt ggt ctg tcc cgt 2060 Thr Tyr Lys Arg Arg Leu Ser Ala Leu Gly Pro Gly Gly Leu Ser Arg 440 445 450 gag cgc gcc ggc atc gag gtt cga gac gtt cac cca tct cac tac ggc 2108 Glu Arg Ala Gly Ile Glu Val Arg Asp Val His Pro Ser His Tyr Gly 455 460 465 cgt atg tgc cca att gag act ccg gaa ggt cca aac att ggc ctg atc 2156 Arg Met Cys Pro Ile Glu Thr Pro Glu Gly Pro Asn Ile Gly Leu Ile 470 475 480 485 ggt tcc ttg gct tcc tat gct cga gtg aac cca ttc ggt ttc att gag 2204 Gly Ser Leu Ala Ser Tyr Ala Arg Val Asn Pro Phe Gly Phe Ile Glu 490 495 500 acc cca tac cgt cgc atc atc gac ggc aag ctg acc gac cag att gac 2252 Thr Pro Tyr Arg Arg Ile Ile Asp Gly Lys Leu Thr Asp Gln Ile Asp 505 510 515 tac ctt acc gct gat gag gaa gac cgc ttc gtt gtt gcg cag gca aac 2300 Tyr Leu Thr Ala Asp Glu Glu Asp Arg Phe Val Val Ala Gln Ala Asn 520 525 530 acg cac tac gac gaa gag ggc aac atc acc gat gag acc gtc act gtt 2348 Thr His Tyr Asp Glu Glu Gly Asn Ile Thr Asp Glu Thr Val Thr Val 535 540 545 cgt ctg aag gac ggc gac atc gcc atg gtt ggc cgc aac gcg gtt gat 2396 Arg Leu Lys Asp Gly Asp Ile Ala Met Val Gly Arg Asn Ala Val Asp 550 555 560 565 tac atg gac gtt tcc cct cgt cag atg gtt tct gtt ggt acc gcg atg 2444 Tyr Met Asp Val Ser Pro Arg Gln Met Val Ser Val Gly Thr Ala Met 570 575 580 att cca ttc ctg gag cac gac gat gct aac cgt gca ctg atg ggc gcg 2492 Ile Pro Phe Leu Glu His Asp Asp Ala Asn Arg Ala Leu Met Gly Ala 585 590 595 aac atg cag aag cag gct gtg cca ctg att cgt gcc gag gct cct ttc 2540 Asn Met Gln Lys Gln Ala Val Pro Leu Ile Arg Ala Glu Ala Pro Phe 600 605 610 gtg ggc acc ggt atg gag cag cgc gca gca tac gac gcc ggc gac ctg 2588 Val Gly Thr Gly Met Glu Gln Arg Ala Ala Tyr Asp Ala Gly Asp Leu 615 620 625 gtt att acc cca gtc gca ggt gtg gtg gaa aac gtt tca gct gac ttc 2636 Val Ile Thr Pro Val Ala Gly Val Val Glu Asn Val Ser Ala Asp Phe 630 635 640 645 atc acc atc atg gct gat gac ggc aag cgc gaa acc tac ctg ctg cgt 2684 Ile Thr Ile Met Ala Asp Asp Gly Lys Arg Glu Thr Tyr Leu Leu Arg 650 655 660 aag ttc cag cgc acc aac cag ggc acc agc tac aac cag aag cct ttg 2732 Lys Phe Gln Arg Thr Asn Gln Gly Thr Ser Tyr Asn Gln Lys Pro Leu 665 670 675 gtt aac ttg ggc gag cgc gtt gaa gct ggc cag gtt att gct gat ggt 2780 Val Asn Leu Gly Glu Arg Val Glu Ala Gly Gln Val Ile Ala Asp Gly 680 685 690 cca ggt acc ttc aat ggt gaa atg tcc ctt ggc cgt aac ctt ctg gtt 2828 Pro Gly Thr Phe Asn Gly Glu Met Ser Leu Gly Arg Asn Leu Leu Val 695 700 705 gcg ttc atg cct tgg gaa ggc cac aac tac gag gat gcg atc atc ctc 2876 Ala Phe Met Pro Trp Glu Gly His Asn Tyr Glu Asp Ala Ile Ile Leu 710 715 720 725 aac cag aac atc gtt gag cag gac atc ttg acc tcg atc cac atc gag 2924 Asn Gln Asn Ile Val Glu Gln Asp Ile Leu Thr Ser Ile His Ile Glu 730 735 740 gag cac gag atc gat gcc cgc gac act aag ctt ggc gcc gaa gaa atc 2972 Glu His Glu Ile Asp Ala Arg Asp Thr Lys Leu Gly Ala Glu Glu Ile 745 750 755 acc cgc gac atc cct aat gtg tct gaa gaa gtc ctc aag gac ctc gac 3020 Thr Arg Asp Ile Pro Asn Val Ser Glu Glu Val Leu Lys Asp Leu Asp 760 765 770 gac cgc ggt att gtc cgc atc ggt gct gat gtt cgt gac ggc gac atc 3068 Asp Arg Gly Ile Val Arg Ile Gly Ala Asp Val Arg Asp Gly Asp Ile 775 780 785 ctg gtc ggt aag gtc acc cct aag ggc gag acc gag ctc acc ccg gaa 3116 Leu Val Gly Lys Val Thr Pro Lys Gly Glu Thr Glu Leu Thr Pro Glu 790 795 800 805 gag cgc ttg ctg cgc gca atc ttc ggt gag aag gcc cgc gaa gtt cgc 3164 Glu Arg Leu Leu Arg Ala Ile Phe Gly Glu Lys Ala Arg Glu Val Arg 810 815 820 gat acc tcc atg aag gtg cct cac ggt gag acc ggc aag gtc atc ggc 3212 Asp Thr Ser Met Lys Val Pro His Gly Glu Thr Gly Lys Val Ile Gly 825 830 835 gtg cgt cac ttc tcc cgc gag gac gac gac gat ctg gct cct ggc gtc 3260 Val Arg His Phe Ser Arg Glu Asp Asp Asp Asp Leu Ala Pro Gly Val 840 845 850 aac gag atg atc cgt atc tac gtt gct cag aag cgt aag atc cag gac 3308 Asn Glu Met Ile Arg Ile Tyr Val Ala Gln Lys Arg Lys Ile Gln Asp 855 860 865 ggc gat aag ctc gct ggc cgc cac ggt aac aag ggt gtt gtc ggt aaa 3356 Gly Asp Lys Leu Ala Gly Arg His Gly Asn Lys Gly Val Val Gly Lys 870 875 880 885 att ttg cct cag gaa gat atg cca ttc ctt cca gac ggc act cct gtt 3404 Ile Leu Pro Gln Glu Asp Met Pro Phe Leu Pro Asp Gly Thr Pro Val 890 895 900 gac atc atc ttg aac acc cac ggt gtt cca cgt cgt atg aac att ggt 3452 Asp Ile Ile Leu Asn Thr His Gly Val Pro Arg Arg Met Asn Ile Gly 905 910 915 cag gtt ctt gag acc cac ctt ggc tgg ctg gca tct gct ggt tgg tcc 3500 Gln Val Leu Glu Thr His Leu Gly Trp Leu Ala Ser Ala Gly Trp Ser 920 925 930 gtg gat cct gaa gat cct gag aac gct gag ctc gtc aag act ctg cct 3548 Val Asp Pro Glu Asp Pro Glu Asn Ala Glu Leu Val Lys Thr Leu Pro 935 940 945 gca gac ctc ctc gag gtt cct gct ggt tcc ttg act gca act cct gtg 3596 Ala Asp Leu Leu Glu Val Pro Ala Gly Ser Leu Thr Ala Thr Pro Val 950 955 960 965 ttc gac ggt gcg tca aac gaa gag ctc gca ggc ctg ctc gct aat tca 3644 Phe Asp Gly Ala Ser Asn Glu Glu Leu Ala Gly Leu Leu Ala Asn Ser 970 975 980 cgt cca aac cgc gac ggc gac gtc atg gtt aac gcg gat ggt aaa gca 3692 Arg Pro Asn Arg Asp Gly Asp Val Met Val Asn Ala Asp Gly Lys Ala 985 990 995 acg ctt atc gac ggt cgc tcc ggt gag cct tac ccg tac ccg gtt 3737 Thr Leu Ile Asp Gly Arg Ser Gly Glu Pro Tyr Pro Tyr Pro Val 1000 1005 1010 tcc atc ggc tac atg tac atg ctg aag ctg cac cac ctc gtt gac 3782 Ser Ile Gly Tyr Met Tyr Met Leu Lys Leu His His Leu Val Asp 1015 1020 1025 gag aag atc cac gca cgt tcc act ggt cct tac tcc atg att acc 3827 Glu Lys Ile His Ala Arg Ser Thr Gly Pro Tyr Ser Met Ile Thr 1030 1035 1040 cag cag cca ctg ggt ggt aaa gca cag ttc ggt gga cag cgt ttc 3872 Gln Gln Pro Leu Gly Gly Lys Ala Gln Phe Gly Gly Gln Arg Phe 1045 1050 1055 ggc gaa atg gag gtg tgg gca atg cag gca tac ggc gct gcc tac 3917 Gly Glu Met Glu Val Trp Ala Met Gln Ala Tyr Gly Ala Ala Tyr 1060 1065 1070 aca ctt cag gag ctg ctg acc atc aag tct gat gac gtg gtt ggc 3962 Thr Leu Gln Glu Leu Leu Thr Ile Lys Ser Asp Asp Val Val Gly 1075 1080 1085 cgt gtc aag gtc tac gaa gca att gtg aag ggc gag aac atc ccg 4007 Arg Val Lys Val Tyr Glu Ala Ile Val Lys Gly Glu Asn Ile Pro 1090 1095 1100 gat cca ggt att cct gag tcc ttc aag gtt ctc ctc aag gag ctc 4052 Asp Pro Gly Ile Pro Glu Ser Phe Lys Val Leu Leu Lys Glu Leu 1105 1110 1115 cag tcc ttg tgc ctg aac gtg gag gtt ctc tcc gca gac ggc act 4097 Gln Ser Leu Cys Leu Asn Val Glu Val Leu Ser Ala Asp Gly Thr 1120 1125 1130 cca atg gag ctc gcg ggt gac gac gac gac ttc gat cag gca ggc 4142 Pro Met Glu Leu Ala Gly Asp Asp Asp Asp Phe Asp Gln Ala Gly 1135 1140 1145 gcc tca ctt ggc atc aac ctg tcc cgt gac gag cgt tcc gac gcc 4187 Ala Ser Leu Gly Ile Asn Leu Ser Arg Asp Glu Arg Ser Asp Ala 1150 1155 1160 gac acc gca tagcagatca gaaaacaacc gctagaaatc aagccataca 4236 Asp Thr Ala 1165 tcccccggac attgaagaga tgttctgggg ggaaagggag ttttacgtgc tcgacgtaaa 4296 cgtcttcgat gagctccgca tcggcctggc caccgccgac gacatccgcc gttggtccaa 4356 gggtgaggtc aagaagccgg agaccatcaa ctaccgaacc ctcaagcctg agaaggacgg 4416 tctgttctgc gagcgtatct tcggtccaac tcgcgactgg gagtgcgcct gcggtaagta 4476 caagcgtgtc cgctacaagg gcatcatctg tgaacgctgt ggcgttgagg tcaccaagtc 4536 caaggtgcgc cgtgagcgca tgggacacat tgagctcgct gcaccagtaa cccacatttg 4596 gtacttcaag ggcgttccat cacgcctcgg ctaccttttg gaccttgctc caaaggacct 4656 ggacctcatc atctacttcg gtgcgaacat catcaccagc gtggacgaag aggctcgcca 4716 cagcgaccag accactcttg aggcagaaat gcttctggag aagaaggacg ttgaggcaga 4776 cgcagagtct gacattgctg agcgtgctga aaagctcgaa gaggatcttg ctgaacttga 4836 ggcagctggc gctaaggccg acgctcgccg caaggttcag gctgctgccg ataaggaaat 4896 gcagcacatc cgtgagcgtg cacagcgcga aatcgatcgt ctcgatgagg tctggcagac 4956 cttcatcaag cttgctccaa agcagatgat ccgcgatgag aagctctacg atgaactgat 5016 cgaccgctac gaggattact tcaccggtgg tatgggtgca gagtccattg aggctttgat 5076 ccagaacttc gaccttgatg ctg 5099 6 1165 PRT Corynebacterium glutamicum 6 Val Leu Glu Gly Pro Ile Leu Ala Val Ser Arg Gln Thr Lys Ser Val 1 5 10 15 Val Asp Ile Pro Gly Ala Pro Gln Arg Tyr Ser Phe Ala Lys Val Ser 20 25 30 Ala Pro Ile Glu Val Pro Gly Leu Leu Asp Leu Gln Leu Asp Ser Tyr 35 40 45 Ser Trp Leu Ile Gly Thr Pro Glu Trp Arg Ala Arg Gln Lys Glu Glu 50 55 60 Phe Gly Glu Gly Ala Arg Val Thr Ser Gly Leu Glu Asn Ile Leu Glu 65 70 75 80 Glu Leu Ser Pro Ile Gln Asp Tyr Ser Gly Asn Met Ser Leu Ser Leu 85 90 95 Ser Glu Pro Arg Phe Glu Asp Val Lys Asn Thr Ile Asp Glu Ala Lys 100 105 110 Glu Lys Asp Ile Asn Tyr Ala Ala Pro Leu Tyr Val Thr Ala Glu Phe 115 120 125 Val Asn Asn Thr Thr Gly Glu Ile Lys Ser Gln Thr Val Phe Ile Gly 130 135 140 Asp Phe Pro Met Met Thr Asp Lys Gly Thr Phe Ile Ile Asn Gly Thr 145 150 155 160 Glu Arg Val Val Val Ser Gln Leu Val Arg Ser Pro Gly Val Tyr Phe 165 170 175 Asp Gln Thr Ile Asp Lys Ser Thr Glu Arg Pro Leu His Ala Val Lys 180 185 190 Val Ile Pro Ser Arg Gly Ala Trp Leu Glu Phe Asp Val Asp Lys Arg 195 200 205 Asp Ser Val Gly Val Arg Ile Asp Arg Lys Arg Arg Gln Pro Val Thr 210 215 220 Val Leu Leu Lys Ala Leu Gly Trp Thr Thr Glu Gln Ile Thr Glu Arg 225 230 235 240 Phe Gly Phe Ser Glu Ile Met Met Ser Thr Leu Glu Ser Asp Gly Val 245 250 255 Ala Asn Thr Asp Glu Ala Leu Leu Glu Ile Tyr Arg Lys Gln Arg Pro 260 265 270 Gly Glu Gln Pro Thr Arg Asp Leu Ala Gln Ser Leu Leu Asp Asn Ser 275 280 285 Phe Phe Arg Ala Lys Arg Tyr Asp Leu Ala Arg Val Gly Arg Tyr Lys 290 295 300 Ile Asn Arg Lys Leu Gly Leu Gly Gly Asp His Asp Gly Leu Met Thr 305 310 315 320 Leu Thr Glu Glu Asp Ile Ala Thr Thr Ile Glu Tyr Leu Val Arg Leu 325 330 335 His Ala Gly Glu Arg Val Met Thr Ser Pro Asn Gly Glu Glu Ile Pro 340 345 350 Val Glu Thr Asp Asp Ile Asp His Phe Gly Asn Arg Arg Leu Arg Thr 355 360 365 Val Gly Glu Leu Ile Gln Asn Gln Val Arg Val Gly Leu Ser Arg Met 370 375 380 Glu Arg Val Val Arg Glu Arg Met Thr Thr Gln Asp Ala Glu Ser Ile 385 390 395 400 Thr Pro Thr Ser Leu Ile Asn Val Arg Pro Val Ser Ala Ala Ile Arg 405 410 415 Glu Phe Phe Gly Thr Ser Gln Leu Ser Gln Phe Met Asp Gln Asn Asn 420 425 430 Ser Leu Ser Gly Leu Thr Tyr Lys Arg Arg Leu Ser Ala Leu Gly Pro 435 440 445 Gly Gly Leu Ser Arg Glu Arg Ala Gly Ile Glu Val Arg Asp Val His 450 455 460 Pro Ser His Tyr Gly Arg Met Cys Pro Ile Glu Thr Pro Glu Gly Pro 465 470 475 480 Asn Ile Gly Leu Ile Gly Ser Leu Ala Ser Tyr Ala Arg Val Asn Pro 485 490 495 Phe Gly Phe Ile Glu Thr Pro Tyr Arg Arg Ile Ile Asp Gly Lys Leu 500 505 510 Thr Asp Gln Ile Asp Tyr Leu Thr Ala Asp Glu Glu Asp Arg Phe Val 515 520 525 Val Ala Gln Ala Asn Thr His Tyr Asp Glu Glu Gly Asn Ile Thr Asp 530 535 540 Glu Thr Val Thr Val Arg Leu Lys Asp Gly Asp Ile Ala Met Val Gly 545 550 555 560 Arg Asn Ala Val Asp Tyr Met Asp Val Ser Pro Arg Gln Met Val Ser 565 570 575 Val Gly Thr Ala Met Ile Pro Phe Leu Glu His Asp Asp Ala Asn Arg 580 585 590 Ala Leu Met Gly Ala Asn Met Gln Lys Gln Ala Val Pro Leu Ile Arg 595 600 605 Ala Glu Ala Pro Phe Val Gly Thr Gly Met Glu Gln Arg Ala Ala Tyr 610 615 620 Asp Ala Gly Asp Leu Val Ile Thr Pro Val Ala Gly Val Val Glu Asn 625 630 635 640 Val Ser Ala Asp Phe Ile Thr Ile Met Ala Asp Asp Gly Lys Arg Glu 645 650 655 Thr Tyr Leu Leu Arg Lys Phe Gln Arg Thr Asn Gln Gly Thr Ser Tyr 660 665 670 Asn Gln Lys Pro Leu Val Asn Leu Gly Glu Arg Val Glu Ala Gly Gln 675 680 685 Val Ile Ala Asp Gly Pro Gly Thr Phe Asn Gly Glu Met Ser Leu Gly 690 695 700 Arg Asn Leu Leu Val Ala Phe Met Pro Trp Glu Gly His Asn Tyr Glu 705 710 715 720 Asp Ala Ile Ile Leu Asn Gln Asn Ile Val Glu Gln Asp Ile Leu Thr 725 730 735 Ser Ile His Ile Glu Glu His Glu Ile Asp Ala Arg Asp Thr Lys Leu 740 745 750 Gly Ala Glu Glu Ile Thr Arg Asp Ile Pro Asn Val Ser Glu Glu Val 755 760 765 Leu Lys Asp Leu Asp Asp Arg Gly Ile Val Arg Ile Gly Ala Asp Val 770 775 780 Arg Asp Gly Asp Ile Leu Val Gly Lys Val Thr Pro Lys Gly Glu Thr 785 790 795 800 Glu Leu Thr Pro Glu Glu Arg Leu Leu Arg Ala Ile Phe Gly Glu Lys 805 810 815 Ala Arg Glu Val Arg Asp Thr Ser Met Lys Val Pro His Gly Glu Thr 820 825 830 Gly Lys Val Ile Gly Val Arg His Phe Ser Arg Glu Asp Asp Asp Asp 835 840 845 Leu Ala Pro Gly Val Asn Glu Met Ile Arg Ile Tyr Val Ala Gln Lys 850 855 860 Arg Lys Ile Gln Asp Gly Asp Lys Leu Ala Gly Arg His Gly Asn Lys 865 870 875 880 Gly Val Val Gly Lys Ile Leu Pro Gln Glu Asp Met Pro Phe Leu Pro 885 890 895 Asp Gly Thr Pro Val Asp Ile Ile Leu Asn Thr His Gly Val Pro Arg 900 905 910 Arg Met Asn Ile Gly Gln Val Leu Glu Thr His Leu Gly Trp Leu Ala 915 920 925 Ser Ala Gly Trp Ser Val Asp Pro Glu Asp Pro Glu Asn Ala Glu Leu 930 935 940 Val Lys Thr Leu Pro Ala Asp Leu Leu Glu Val Pro Ala Gly Ser Leu 945 950 955 960 Thr Ala Thr Pro Val Phe Asp Gly Ala Ser Asn Glu Glu Leu Ala Gly 965 970 975 Leu Leu Ala Asn Ser Arg Pro Asn Arg Asp Gly Asp Val Met Val Asn 980 985 990 Ala Asp Gly Lys Ala Thr Leu Ile Asp Gly Arg Ser Gly Glu Pro Tyr 995 1000 1005 Pro Tyr Pro Val Ser Ile Gly Tyr Met Tyr Met Leu Lys Leu His 1010 1015 1020 His Leu Val Asp Glu Lys Ile His Ala Arg Ser Thr Gly Pro Tyr 1025 1030 1035 Ser Met Ile Thr Gln Gln Pro Leu Gly Gly Lys Ala Gln Phe Gly 1040 1045 1050 Gly Gln Arg Phe Gly Glu Met Glu Val Trp Ala Met Gln Ala Tyr 1055 1060 1065 Gly Ala Ala Tyr Thr Leu Gln Glu Leu Leu Thr Ile Lys Ser Asp 1070 1075 1080 Asp Val Val Gly Arg Val Lys Val Tyr Glu Ala Ile Val Lys Gly 1085 1090 1095 Glu Asn Ile Pro Asp Pro Gly Ile Pro Glu Ser Phe Lys Val Leu 1100 1105 1110 Leu Lys Glu Leu Gln Ser Leu Cys Leu Asn Val Glu Val Leu Ser 1115 1120 1125 Ala Asp Gly Thr Pro Met Glu Leu Ala Gly Asp Asp Asp Asp Phe 1130 1135 1140 Asp Gln Ala Gly Ala Ser Leu Gly Ile Asn Leu Ser Arg Asp Glu 1145 1150 1155 Arg Ser Asp Ala Asp Thr Ala 1160 1165 7 1775 DNA Corynebacterium glutamicum CDS (500)..(880) 7 cagctctaca agagtgtcta agtggcgggc attccatgct ttggaggagc gatcttcaaa 60 ttcctccaaa gtgagttgac ctcgggaaac agctgcagaa agttcatcca cgacttggtt 120 tcggttaagg tcagtggcga gcttctttgc tggttcgttt ccttgaggaa cagtcatggg 180 aaccattcta acaagggatt tggtgttttc tgcggctagc tgataatgtg aacggctgag 240 tcccactctt gtagttggga attgacggca cctcgcactc aagcgcggta tcgcccctgg 300 ttttccggga cgcggtggcg catgtttgca tttgatgagg ttgtccgtga catgtttggt 360 cgggccccaa aaagagcccc cttttttgcg tgtctggaca ctttttcaaa tccttcgcca 420 tcgacaagct cagccttcgt gttcgtcccc cgggcgtcac gtcagcagtt aaagaacaac 480 tccgaaataa ggatggttc atg cca act att cag cag ctg gtc cgt aag ggc 532 Met Pro Thr Ile Gln Gln Leu Val Arg Lys Gly 1 5 10 cgc cac gat aag tcc gcc aag gtg gct acc gcg gca ctg aag ggt tcc 580 Arg His Asp Lys Ser Ala Lys Val Ala Thr Ala Ala Leu Lys Gly Ser 15 20 25 cct cag cgt cgt ggc gta tgc acc cgt gtg tac acc acc acc cct aag 628 Pro Gln Arg Arg Gly Val Cys Thr Arg Val Tyr Thr Thr Thr Pro Lys 30 35 40 aag cct aac tct gct ctt cgt aag gtc gct cgt gtg cgc ctt acc tcc 676 Lys Pro Asn Ser Ala Leu Arg Lys Val Ala Arg Val Arg Leu Thr Ser 45 50 55 ggc atc gag gtt tcc gct tac atc cct ggt gag ggc cac aac ctg cag 724 Gly Ile Glu Val Ser Ala Tyr Ile Pro Gly Glu Gly His Asn Leu Gln 60 65 70 75 gag cac tcc atg gtg ctc gtt cgc ggt ggt cgt gtt aag gac ctc cca 772 Glu His Ser Met Val Leu Val Arg Gly Gly Arg Val Lys Asp Leu Pro 80 85 90 ggt gtc cgt tac aag atc gtc cgt ggc gca ctg gat acc cag ggt gtt 820 Gly Val Arg Tyr Lys Ile Val Arg Gly Ala Leu Asp Thr Gln Gly Val 95 100 105 aag gac cgc aag cag gct cgt tcc ccg cta cgg cgc gaa gag ggg ata 868 Lys Asp Arg Lys Gln Ala Arg Ser Pro Leu Arg Arg Glu Glu Gly Ile 110 115 120 att aaa aat gcg taaatcagca gctcctaagc gtccagtagt tcaggaccct 920 Ile Lys Asn Ala 125 gtatacaagt ccgagctcgt tacccagctc gtaaacaaga tcctcatcgg tggcaagaag 980 tccaccgcag agcgcatcgt ctacggtgca ctcgagatct gccgtgagaa gaccggcacc 1040 gatccagtag gaaccctcga gaaggctctc ggcaacgtgc gtccagacct cgaagttcgt 1100 tcccgccgtg ttggtggcgc tacctaccag gtgccagtgg atgttcgccc agagcgcgca 1160 aacaccctcg cactgcgttg gttggtaacc ttcacccgtc agcgtcgtga gaacaccatg 1220 atcgagcgtc ttgcaaacga acttctggat gcagccaacg gccttggcgc ttccgtgaag 1280 cgtcgcgaag acacccacaa gatggcagag gccaaccgcg ccttcgctca ctaccgctgg 1340 tagtactgcc aagacatgaa agcccaatca cctttaagat caacgcctgc cggcgccctt 1400 cacatttgaa taagctggca gcctgcgttt cttcaaggcg actgggcttt tagtctcatt 1460 aatgcagttc accgctgtaa gatagctaaa tagaaacact gtttcggcag tgtgttacta 1520 aaaaatccat gtcacttgcc tcgagcgtgc tgcttgaatc gcaagttagt ggcaaaatgt 1580 aacaagagaa ttatccgtag gtgacaaact ttttaatact tgggtatctg tcatggatac 1640 cccggtaata aataagtgaa ttaccgtaac caacaagttg gggtaccact gtggcacaag 1700 aagtgcttaa ggatctaaac aaggtccgca acatcggcat catggcgcac atcgatgctg 1760 gtaagaccac gacca 1775 8 127 PRT Corynebacterium glutamicum 8 Met Pro Thr Ile Gln Gln Leu Val Arg Lys Gly Arg His Asp Lys Ser 1 5 10 15 Ala Lys Val Ala Thr Ala Ala Leu Lys Gly Ser Pro Gln Arg Arg Gly 20 25 30 Val Cys Thr Arg Val Tyr Thr Thr Thr Pro Lys Lys Pro Asn Ser Ala 35 40 45 Leu Arg Lys Val Ala Arg Val Arg Leu Thr Ser Gly Ile Glu Val Ser 50 55 60 Ala Tyr Ile Pro Gly Glu Gly His Asn Leu Gln Glu His Ser Met Val 65 70 75 80 Leu Val Arg Gly Gly Arg Val Lys Asp Leu Pro Gly Val Arg Tyr Lys 85 90 95 Ile Val Arg Gly Ala Leu Asp Thr Gln Gly Val Lys Asp Arg Lys Gln 100 105 110 Ala Arg Ser Pro Leu Arg Arg Glu Glu Gly Ile Ile Lys Asn Ala 115 120 125 9 24 DNA ARTIFICIAL SEQUENCE SYNTHETIC DNA 9 acaatgtgac tcgtgatttt tggg 24 10 20 DNA ARTIFICIAL SEQUENCE SYNTHETIC DNA 10 ggaaacgtcc atgtaatcaa 20 11 20 DNA ARTIFICIAL SEQUENCE SYNTHETIC DNA 11 aacacgcact acgacgaaga 20 12 20 DNA ARTIFICIAL SEQUENCE SYNTHETIC DNA 12 cagcatcaag gtcgaagttc 20 

1. An isolated polynucleotide which encodes a protein comprising the amino acid sequence of SEQ ID NO:2.
 2. The isolated polynucleotide of claim 1, wherein said protein has the activity of the β-subunit of RNA polymerase B.
 3. An isolated polynucleotide which comprises SEQ ID NO:1.
 4. An isolated polynucleotide which is complimentary to the polynucleotide of claim
 3. 5. An isolated polynucleotide which is at least 70% identical to the polynucleotide of claim
 3. 6. An isolated polynucleotide which is at least 80% identical to the polynucleotide of claim
 3. 7. An isolated polynucleotide which is at least 90% identical to the polynucleotide of claim
 3. 8. An isolated polynucleotide which hybridizes under stringent conditions to the polynucleotide of claim 3; wherein said stringent conditions comprise washing in 5×SSC at a temperature from 50 to 68° C.
 9. The isolated polynucleotide of claim 3, which encodes a protein having the activity of the β-subunit of RNA polymerase B.
 10. An isolated polynucleotide which comprises at least 15 consecutive nucleotides of the polynucleotide of claim
 3. 11. An isolated polypeptide which comprises the amino acid sequence of SEQ ID NO:2.
 12. An isolated polypeptide which comprises the amino acid sequence of SEQ ID NO:4.
 13. An isolated polypeptide which comprises the amino acid sequence of SEQ ID NO:6.
 14. An isolated polynucleotide which encodes a protein comprising the amino acid sequence of SEQ ID NO:4.
 15. An isolated polynucleotide which comprises SEQ ID NO:3.
 16. An isolated polynucleotide which encodes a protein comprising the amino acid sequence of SEQ ID NO:6.
 17. An isolated polynucleotide which comprises SEQ ID NO:5.
 18. A vector comprising the isolated polynucleotide of claim
 1. 19. A vector comprising the isolated polynucleotide of claim
 3. 20. A vector comprising the isolated polynucleotide of claim
 14. 21. A vector comprising the isolated polynucleotide of claim
 15. 22. A vector comprising the isolated polynucleotide of claim
 16. 23. A vector comprising the isolated polynucleotide of claim
 17. 24. A host cell comprising the isolated polynucleotide of claim
 1. 25. A host cell comprising the isolated polynucleotide of claim
 3. 26. A host cell comprising the isolated polynucleotide of claim
 14. 27. A host cell comprising the isolated polynucleotide of claim
 15. 28. A host cell comprising the isolated polynucleotide of claim
 16. 29. A host cell comprising the isolated polynucleotide of claim
 17. 30. The host cell of claim 24, which is a Coryneform bacterium.
 31. The host cell of claim 25, which is a Coryneform bacterium.
 32. The host cell of claim 26, which is a Coryneform bacterium.
 33. The host cell of claim 27, which is a Coryneform bacterium.
 34. The host cell of claim 28, which is a Coryneform bacterium.
 35. The host cell of claim 29, which is a Coryneform bacterium.
 36. The host cell of claim 24, wherein said host cell is selected from the group consisting of Coryneform glutamicum, Corynebacterium acetoglutamicum, Corynebacterium acetoacidophilum, Corynebacterium thermoaminogenes, Corynebacterium melassecola, Brevibacterium flavum, Brevibacterium lactofermentum, and Brevibacterium divaricatum.
 37. The host cell of claim 24, wherein said host cell is selected from the group consisting of Corynebacterium glutamicum FERM 1709, Brevibacterium flavum FERM-P 1708, Brevibacterium lactofermentum FERM-P1712, Corynebacterium glutamicum FERM-P6463, Corynebacterium glutamicum FERM-P6464, Corynebacterium glutamicum DM58-1, Corynebacterium glutamicum DG 52-5, Corynebacterium glutamicum DSM 5714 and Corynebacterium glutamicum DSM-12866.
 38. The host cell of claim 25, wherein said host cell is selected from the group consisting of Coryneform glutamicum, Corynebacterium acetoglutamicum, Corynebacterium acetoacidophilum, Corynebacterium thermoaminogenes, Corynebacterium melassecola, Brevibacterium flavum, Brevibacterium lactofermentum, and Brevibacterium divaricatum.
 39. The host cell of claim 25, wherein said host cell is selected from the group consisting of Corynebacterium glutamicum FERM 1709, Brevibacterium flavum FERM-P 1708, Brevibacterium.lactofermentum FERM-P1712, Corynebacterium glutamicum FERM-P6463, Corynebacterium glutamicum FERM-P6464, Corynebacterium glutamicum DM58-1, Corynebacterium glutamicum DG 52-5, Corynebacterium glutamicum DSM 5714 and Corynebacterium glutamicum DSM-12866.
 40. A Coryneform bacterium which comprises an enhanced rpoB gene.
 41. The Coryneform bacterium of claim 40, wherein said rpoB gene comprises the polynucleotide sequence of SEQ ID NO:1.
 42. The Coryneform bacterium of claim 40, wherein said enhanced rpoB gene comprises the polynucleotide sequence of SEQ ID NO:3.
 43. The Coryneform bacterium of claim 40, wherein said enhanced rpoB gene comprises the polynucleotide sequence of SEQ ID NO:5.
 44. Coryneform glutamicum DSM
 13993. 45. Coryneform glutamicum DSM
 13994. 46. A process for producing L-amino acids comprising culturing the host cell of claim 24 in a medium suitable for the expression of the polynucleotide; and collecting the L-amino acid.
 47. The process of claim 46, wherein said L-amino acid is L-lysine or L-glutamate.
 48. The process of claim 46, wherein said L-amino acid is L-lysine and the host cell further comprises at least one gene whose expression is enhanced, wherein said gene is selected from the group consisting of dapA, gap, tpi, pgk, zwf, pyc, mqo, lys C, lys E, zwa1 and rpsL.
 49. The process of claim 46, wherein the host cell further comprises at least one gene whose expression is attenuated, wherein said gene is selected from the group consisting of pck gene, pgi gene, poxB, and zwa2.
 50. A process for producing L-amino acids comprising culturing the host cell of claim 25 in a medium suitable for the expression of the polynucleotide; and collecting the L-amino acid.
 51. The process of claim 50, wherein said L-amino acid is L-lysine or L-glutamate.
 52. The process of claim 50, wherein wherein said L-amino acid is L-lysine and the host cell further comprises at least one gene whose expression is enhanced, wherein said gene is selected from the group consisting of dapA, gap, tpi, pgk, zwf, pyc, mqo, lys C, lys E, zwa1 and rpsL.
 53. The process of claim 50, wherein the host cell further comprises at least one gene whose expression is attenuated, wherein said gene is selected from the group consisting of pck gene, pgi gene, poxB, and zwa2.
 54. A process for producing L-amino acids comprising culturing the host cell of claim 26 in a medium suitable for the expression of the polynucleotide; and collecting the L-amino acid.
 55. The process of claim 54, wherein said L-amino acid is L-lysine or L-glutamate.
 56. The process of claim 54, wherein wherein said L-amino acid is L-lysine and the host cell further comprisesat least one gene whose expression is enhanced, wherein said gene is selected from the group consisting of dapA, gap, tpi, pgk, zwf, pyc, mqo, lys C, lys E, zwa1 and rpsL.
 57. The process of claim 54, wherein the host cell further comprises at least one gene whose expression is attenuated, wherein said gene is selected from the group consisting of pck gene, pgi gene, poxB, and zwa2.
 58. A process for producing L-amino acids comprising culturing the host cell of claim 26 in a medium suitable for the expression of the polynucleotide; and collecting the L-amino acid.
 59. The process of claim 58, wherein said L-amino acid is L-lysine or L-glutamate.
 60. The process of claim 58, wherein wherein said L-amino acid is L-lysine and the host cell further comprisesat least one gene whose expression is enhanced, wherein said gene is selected from the group consisting of dapA, gap, tpi, pgk, zwf, pyc, mqo, lys C, lys E, zwa1 and rpsL.
 61. The process of claim 58, wherein the host cell further comprises at least one gene whose expression is attenuated, wherein said gene is selected from the group consisting of pck gene, pgi gene, poxB, and zwa2.
 62. A process for producing L-amino acids comprising culturing the host cell of claim 27 in a medium suitable for the expression of the polynucleotide; and collecting the L-amino acid.
 63. The process of claim 62, wherein said L-amino acid is L-lysine or L-glutamate.
 64. The process of claim 62, wherein wherein said L-amino acid is L-lysine and the host cell further comprisesat least one gene whose expression is enhanced, wherein said gene is selected from the group consisting of dapA, gap, tpi, pgk, zwf, pyc, mqo, lys C, lys E, zwa1 and rpsL.
 65. The process of claim 62, wherein the host cell further comprises at least one gene whose expression is attenuated, wherein said gene is selected from the group consisting of pck gene, pgi gene, poxB, and zwa2.
 66. A process for producing L-amino acids comprising culturing the host cell of claim 28 in a medium suitable for the expression of the polynucleotide; and collecting the L-amino acid.
 67. The process of claim 66, wherein said L-amino acid is L-lysine or L-glutamate.
 68. The process of claim 66, wherein wherein said L-amino acid is L-lysine and the host cell further comprisesat least one gene whose expression is enhanced, wherein said gene is selected from the group consisting of dapA, gap, tpi, pgk, zwf, pyc, mqo, lys C, lys E, zwa1 and rpsL.
 69. The process of claim 66, wherein the host cell further comprises at least one gene whose expression is attenuated, wherein said gene is selected from the group consisting of pck gene, pgi gene, poxB, and zwa2.
 70. A process for producing L-amino acids comprising culturing the host cell of claim 29 in a medium suitable for the expression of the polynucleotide; and collecting the L-amino acid.
 71. The process of claim 70, wherein said L-amino acid is L-lysine or L-glutamate.
 72. The process of claim 70, wherein wherein said L-amino acid is L-lysine and the host cell further comprisesat least one gene whose expression is enhanced, wherein said gene is selected from the group consisting of dapA, gap, tpi, pgk, zwf, pyc, mqo, lys C, lys E, zwa1 and rpsL.
 73. The process of claim 70, wherein the host cell further comprises at least one gene whose expression is attenuated, wherein said gene is selected from the group consisting of pck gene, pgi gene, poxB, and zwa2.
 74. A process for screening for polynucleotides which encode a protein having the activity of the β-subunit of RNA polymerase B comprising hybridizing the isolated polynucleotide of claim 1 to the polynucleotide to be screened; expressing the polynucleotide to produce a protein; and detecting the presence or absence of the activity of the β-subunit of RNA polymerase B in said protein.
 75. A process for screening for polynucleotides which encode a protein having the activity of the β-subunit of RNA polymerase B comprising hybridizing the isolated polynucleotide of claim 3 to the polynucleotide to be screened; expressing the polynucleotide to produce a protein; and detecting the presence or absence of the activity of the β-subunit of RNA polymerase B in said protein.
 76. A process for screening for polynucleotides which encode a protein having the activity of the β-subunit of RNA polymerase B comprising hybridizing the isolated polynucleotide of claim 15 to the polynucleotide to be screened; expressing the polynucleotide to produce a protein; and detecting the presence or absence the activity of the β-subunit of RNA polymerase B in said protein.
 77. A process for screening for polynucleotides which encode a protein having the activity of the β-subunit of RNA polymerase B comprising hybridizing the isolated polynucleotide of claim 17 to the polynucleotide to be screened; expressing the polynucleotide to produce a protein; and detecting the presence or absence the activity of the β-subunit of RNA polymerase B in said protein.
 78. A method for detecting a nucleic acid with at least 70% homology to nucleotide of claim 1, comprising contacting a nucleic acid sample with a probe or primer comprising at least 15 consecutive nucleotides of the nucleotide sequence of claim 1, or at least 15 consecutive nucleotides of the complement thereof.
 79. A method for producing a nucleic acid with at least 70% homology to nucleotide of claim 1, comprising contacting a nucleic acid sample with a primer comprising at least 15 consecutive nucleotides of the nucleotide sequence of claim 1, or at least 15 consecutive nucleotides of the complement thereof.
 80. A method for detecting a nucleic acid with at least 70% homology to nucleotide of claim 3, comprising contacting a nucleic acid sample with a probe or primer comprising at least 15 consecutive nucleotides of the nucleotide sequence of claim 3, or at least 15 consecutive nucleotides of the complement thereof.
 81. A method for producing a nucleic acid with at least 70% homology to nucleotide of claim 3, comprising contacting a nucleic acid sample with a primer comprising at least 15 consecutive nucleotides of the nucleotide sequence of claim 3, or at least 15 consecutive nucleotides of the complement thereof.
 82. A method for making a β-subunit of RNA polymerase B, comprising: culturing the host cell of claim 23 for a time and under conditions suitable for expression of the β-subunit of RNA polymerase B, and collecting the β-subunit of RNA polymerase B.
 83. A method for making a β-subunit of RNA polymerase B, comprising: culturing the host cell of claim 24 for a time and under conditions suitable for expression of the β-subunit of RNA polymerase B, and collecting the β-subunit of RNA polymerase B.
 84. A method for making a β-subunit of RNA polymerase B, comprising: culturing the host cell of claim 25 for a time and under conditions suitable for expression of the β-subunit of RNA polymerase B, and collecting the β-subunit of RNA polymerase B.
 85. A method for making a β-subunit of RNA polymerase B, comprising: culturing the host cell of claim 26 for a time and under conditions suitable for expression of the β-subunit of RNA polymerase B, and collecting the β-subunit of RNA polymerase B.
 86. A method for making a β-subunit of RNA polymerase B, comprising: culturing the host cell of claim 27 for a time and under conditions suitable for expression of the β-subunit of RNA polymerase B, and collecting the β-subunit of RNA polymerase B.
 87. A method for making a β-subunit of RNA polymerase B, comprising: culturing the host cell of claim 28 for a time and under conditions suitable for expression of the β-subunit of RNA polymerase B, and collecting the β-subunit of RNA polymerase B. 